Exploring the Binding Site of Ubiquinone in Complex I by EPR/DEER

Exploring the Binding Site of Ubiquinone in Complex I by EPR/DEER
Müge Aksoyoglu*, Udo Glessner**, Thomas Spatzal**, Erik Schleicher*, Stefan Weber*
* Department of Physical Chemistry, Albert-Ludwigs-Universität Freiburg, Germany
** Department of Biochemistry, Albert-Ludwigs-Universität Freiburg, Germany
The proton pumping NADH:ubiquinone oxidoreductase, the respiratory complex I, couples
electron transfer from NADH to ubiquinone with the translocation of protons across the
membrane [1]. Ubiquinone, which binds to complex I, is a key component of the electrontransport
chain because it delivers electrons from complex I to complex III. Recently, the
three-dimensional structure of the entire complex I from Thermus thermophilius was
determined [2]. However, the structure does not show the presence of bound quinone but a
large cavity at the end of the peripheral arm was proposed as binding site. In this study, sitedirected
spin labelling (SDSL) in combination with EPR/DEER spectroscopy is used to
localize the ubiquinone binding site in complex I on a molecular level. Various positions in
the protein as well as decyl-ubiquinone (Q10) were spin labelled with MTSL to measure
distances between the two labels via cw-EPR and DEER experiments. We present some of
our results considering the binding site of ubiquinone.
Moreover, we present preliminary data on the FeMo cofactor from nitrogenase, which reduces
the dinitrogen triple bond under atmospheric pressure and temperature, thereby forming
ammonia in a reaction coupled to the hydrolysis of ATP [3]. Various EPR and HYSCORE
methods were used to investigate the FeMo cofactor, which is not yet fully understood in
terms of its composition.
[1] Pohl, T., Spatzal, T., Aksoyoglu, M., Schleicher, E., Rostas, A.M., Lay, H., Glessner, U.,
Boudon, C., Hellwig, P., Weber, S., Friedrich, T., Spin labelling of the Escherichia coli
NADH ubiquinone oxidoreductase (complex I). Biochim. Biophys. Acta 2010, 1797,
1894–1900.
[2] Efremov, R.G., Baradaran, R., Sazanov, L.A., The architecture of respiratory complex I.
Nature 2010, 465, 441–447.
[3] Lee, H., Benton, P., Laryukhin, M., Igarashi, R., Dean, D.R., Seefeldt, L.C., Hoffmann,
B.M., The interstitial atom of the nitrogenase FeMo-cofactor: ENDOR and ESEEM show
it is not exchangeable nitrogen. J. Am. Chem. Soc. 2003, 125, 5604–5605.
1

PELDOR Study of Nitroxide-Labeled Proteins
Florian Altvater, Stephan Rein, Stefan Weber and Sylwia Kacprzak
Investigating biomacromolecules in solution with pulsed electron-electron double
resonance (PELDOR) spectroscopy yields distance distributions, which provide
information about structure and dynamics. The distances are measured between two
paramagnetic centers, e. g. nitroxides, that are introduced into the desired positions
via site-directed spin labeling. PELDOR is a method of choice especially when no
crystal structure is available. Based on the results of distance measurements initial
model structures can be refined or the conformational changes during enzymatic
actions can be followed.
We present the application of PELDOR method on the examples of two distinct
proteins, Gyrase of Bacillus subtilis and Agp1 of Agrobacterium tumefaciens. Gyrase
is a heterotetramer consisting of two A and two B subunits. Preliminary results of the
distance distribution in the Gyrase A dimmer of the mutant N399C are shown. The
second example present molecular dynamics (MD) simulation approach to refine the
model structure of phytochrome Agp1 of Agrobacterium tumefaciens.
2

A Flexible Approach to Benzotropolones
Deniz Arican and Reinhard Brückner
Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität, Albertstraße 21,
79104 Freiburg, Germany, e-mail: deniz.arican@uranus.uni-freiburg.de
Benzotropolones (1) are bicyclic aromatic compounds, which are found in plants and fungi.
Besides they are interesting from an industrial point of view. Within the last six years patents
have been granted regarding their use as anti-inflammatory agents, anti-obesity medication,
UV-absorbers in sunscreens,
antimicrobial
agents, and for the
stabilization of household,
cosmetic, and
nutritional products.
Perceiving benzotropolone (1) not as a benz-annulated aromatic but rather as the enol
tautomer of 1,2-diketone 5 gave rise to the idea of tracing back the latter to the monoketal 6.
Being a substituted cycloheptenone of sorts 6 should be accessible by a ring-closing
metathesis of diolefin
7. The
latter could
originate from
the aldehyde 8
and the aromatic
compound 9.
Following this strategy benzotropolones with 1-3 substituents at various locations were
synthesized in overall yields from 18% over 8 steps to 78% over 6 steps.
3

Enantiomerically Pure para-Benzoquinone Spiroketals –
Novel Building-Blocks for Stereochemically Versatile
Syntheses of Cyclohexitols and Carbasugars
Johannes Aucktor, a Chiara Anselmi, a,b Reinhard Brückner,* a and Manfred Keller a
a) Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität, Albertstraße 21, 79104 Freiburg, Germany
b) New address: Dipartimento di Chimica, Università degli Studi di Perugia, Via Elce di Sotto 8, 06123 Perugia, Italy
Conduritols (1), quercitols (2), and inositols (3) are structurally diverse polyhydroxy-
cyclohexanes (“cyclohexitols”) from nature. Carbapyranoses (4) are analogously
structured unnatural compounds. The great stereochemical variety of these species
makes them challenging synthetic targets.
HO
1
*
OH
*
* OH
* OH
HO
* * OH
HO * * * OH
2 OH
O
5
HO
*
O
O O *
*
H/R
*
OH
HO
*
OH
*
* OH
HO * * * OH
3 OH
OH
* * OH
HO * * * OH
4 OH
We recognized that (almost) all such compounds may stem from one common pre-
cursor 5. The latter was accessible from hydroquinone with �99% ee. 5 contains a
rigid tricyclic scaffold. It allows to functionalize the cyclohexenone-moiety with a high
degree of diastereocontrol. Well-working functionalizations include cis-vic-dihydroxy-
lation, nucleophilic epoxidation, carbonyl group reduction, and enolate hydroxylation.
Precursors of the above-mentioned polyhydroxycyclohexanes are accessible under a
variety of conditions from such functionalized scaffolds.
4

Biaxial Liquid Crystalline Polymers
F. Brömmel 1 , D. Kramer 1 , P. Benzie 2 , G. Osterwinter 1 , H. Finkelmann 1 , S. Elston 2 , A. Hoffmann 1
1.) Institut für Makromolekulare Chemie, Albert-Ludwigs-Universität Freiburg, 79104 Freiburg
2.) Department of Engineering Science, University of Oxford, Oxford, Parks Road, Oxford, OX1
3PJ, UK
The recent experimental discovery of biaxial liquid crystal
(LC) phases provides the opportunity to develop the technology
for a new generation of faster biaxial LC displays.
These expectations are based on the existence of a secondary
director that can be controlled by external fields.
It is estimated that the reorientation of the secondary director
can be more than one order of magnitude faster
than the reorientation of the uniaxial nematic director.
However, fundamental questions concerning the biaxial
nematic phase behaviour, its alignment with respect to
external fields and its relation to the molecular architecture
are still open.
So far, a large number of nematic polymers and elastomers have been found to form biaxial nematic
phases. We deuterated these systems in various positions. This allowed us to compare the obtained
biaxiality parameters from 2 H NMR experiments on directly deutrated monomers and deuterated probe
molecules, that are commonly used to determine biaxiality in order to overcome the synthetical effort
of deuteration. Also, it is possible to vary the molecular geometry of the mesogens while preserving
the biaxiality. This way the influence of the molecular geometry on biaxiality was studied.
The temperature dependence of the biaxiality for all polymers investigated so far suggests that the
formation of the biaxial phase is connected to the glass transition. Accordingly, the dynamic of the
polymer chains plays a decisive role in the formation of the biaxial phase.
For potential applications, the manipulation of the minor director by external fields without affecting
the orientation of the principal axis is crucial. X-ray scattering experiments and conoscopy measurements
performed under mechanical deformation reveal that the reorientation of the secondary director
occurs before the reorientation of the primary nematic director sets in. This proves the selective addressability
of the biaxial director by mechanical fields.
5

Single molecule spectroscopy of membrane bound H + -ATPsynthases from E. coli
Markus Burger *, Peter Gräber*
* Institut für Physikalische Chemie, Albert-Ludwigs Universität Freiburg, Germany
Markus.Burger@physchem.uni-freiburg.de
Subunit movements within the H + -ATPsynthase from E. coli (EF0F1) are investigated by
single molecule FRET spectroscopy. The enzyme is covalently labeled at cysteines within the
γ-subunit (γC106) and the ε-subunit (εC114). The labeling procedure is carried out with a
mixture of both fluorescent dyes. Therefore, the binding at the two positions with the donorfluorophor
(ATTO532) and the acceptor-fluorophor (ATTO610) occurs statistically. The
donor and acceptor labeled EF0F1 is integrated into liposomes and single-pair fluorescence
resonance energy transfer is measured in freely diffusing proteoliposomes with a confocal two
channel microscope. Addition of AMPPNP (a non-hydrolysable analogue of ATP) leads to a
conformational change of the ε-subunit from an extended to a folded form. Analysis of
ensemble-FRET and single-pair-FRET indicates distance changes in the range of 4 nm. The
fluorescence time traces obtained with single-pair-FRET reveal a time-dependent switching of
the ε-subunit in the presence of nucleotides.
ε114 UP
γ106C
ε114 DOWN
Fig. 1. The γ (blue) and ε subunits from E.coli. The ε subunit is shown in up (cyan-green) and
down (cyan-red) conformation. Labeled positions are marked with spheres.
6
International Bunsen Discussion Meeting – FRET in Life Sciences – 27th - 30th March 2011

Jan Caspar
Arbeitskreis Prof. Dr. P. Spiteller
Institut für Organische Chemie und Biochemie
Titel: Verwundungsaktivierte Verteidigung in Marasmius oreades durch Cyanwasserstoff
Pilze entwickelten wie auch Pflanzen eine Reihe von Abwehrmechanismen, um sich gegen Fraßfeinde
und Schädlinge schützen zu können. Bei der verwundungsaktivierten Verteidigung liegen inaktive
Vorläuferverbindungen der eigentlichen Wirkstoffe vor, welche durch enzymatische Umwandlung im
Verteidigungsfall in ihre bioaktive Form überführt werden. Neben komplexeren, biologisch aktiven
Verbindungen werden auch einfachere Toxine zur Verteidigung eingesetzt. So konnte aus
Fruchtkörpern der Art Aleurodiscus amorphus ein Cyanhydrin isoliert werden, welches durch einen
oxidativen Mechanismus zu HCN und Aleurodiscoester umgesetzt wird. In Fruchtkörpern der Art
Marasmius oreades konnte nach Beschädigung ebenfalls eine Cyanidfreisetzung festgestellt werden.
Die zugehörige Ausgangsverbindung ist jedoch nicht bekannt. Erste Experimente legen nahe, dass es
sich hierbei um ein hochpolares, geschütztes Cyanhydrin handelt. Die Substanz soll mittels HPLC-
Techniken isoliert und NMR-spektroskopisch und massenspektrometrisch vollständig charakterisiert
werden. Da die Substanz kein Chromophor enthält, muss die Detektion durch eine Abbaureaktion zur
Freisetzung von Cyanid sowie anschließendem Cyanidnachweis durchgeführt werden.
7

Crater morphologies in impact experiments into sandstone targets
A.Dufresne, M.H.Poelchau, T.Kenkmann, and the MEMIN-Team
Department of Geological Sciences – Geology, Albert-Ludwigs Universität Freiburg, Germany
Understanding the dynamics of a hypervelocity body impacting onto a planetary surface can
give numerous insights into the processes that formed the solar system. During an impact
process, the impacting projectile initially excavates a roughly hemispherical cavity in the
target, known as the “transient crater”, which is usually modified by late stage processes.
Constraining the dimensions of the transient crater and late stage modification features like
spallation lead to a better understanding of the physical parameters that affect crater
formation. Therefore, impact experiments into sandstone cubes were performed at the two-
stage light gas acceleration facilities of the EMI Freiburg. Spherical projectiles of steel,
meteoritic iron and aluminium were accelerated to velocities of 2.5-8 km/s and impact
energies from 0.7 to 58 kJ. In a detailed study, the processes influencing crater volume,
morphology and spall behaviour were investigated through visual mapping, 3D digital
modelling and 2D profile analyses. All experimental impact craters in dry sandstone targets
share four characteristic morphological features: (1) a light-coloured, fragile, highly
fragmented central part, (2) areas of “arrested spallation” where the target is fractured but the
outlined spall fragments remains within the crater context, (3) typically two inner depressions,
and (4) an outer, shallow (10-20°) dipping area. Constraining the percentage of spallation
within a crater and determining the shape and volume of the transient crater are vital for
comparison with non-brittle materials like metals, with numerical modelling results, and with
natural impact craters. Transient crater morphologies are derived from (i) parabolas fitted to
the central depressions of crater profiles, (ii) reconstructed spallation dimensions, (iii) ejecta
imprint diameters on catcher systems, (iv) ejecta cone angle and dynamic crater diameter
from high-speed videos, and (i) the distribution of ejecta weight fractions. Results show that
spallation, as a late-stage cratering process, increases the volume of the transient crater by a
factor of 5-10. The refined transient crater model will be applied to other experimental and to
real-size impact craters in due course.
8

in-vivo localization of the OXPHOS complexes in E. coli
Erhardt H. and Friedrich T.
Institute for Organic Chemistry and Biochemistry,
Albert-Ludwigs-University Freiburg, Germany
Biological membranes show a high level of organization of their compounds. Lipids
and membrane proteins are not randomly distributed within these membranes, but
organized in distinct and dynamic clusters and show a huge variety in localization
patterns. It has been proposed that protein complexes connected by a common
substrate chain form supercomplexes, which are located in specific areas within the
membrane. Such an organization provides advantages such as stabilization of the
individual complexes or catalytic enhancement.
The aerobic OXPHOS system of E. coli consists of several multi-subunit enzyme
complexes. To address the question whether these complexes are localized in
distinct lipid areas and if they show any dynamic behavior, we decorated the
membrane protein complexes with different fluorescent proteins and visualized their
distribution in the membrane by in-vivo fluorescence microscopy. FP-decorated
variants of the NADH:ubiquinone oxidoreductase, the succinate dehydrogenase, the
cytochrome-bd complex and the F0F1-ATP-synthase, were created by means of λ-
RED mediated mutagenesis. The labeled complexes were catalytically active and
fully assembled. By fluorescence microscopy we observed an uneven distribution of
the OXPHOS complexes in the cytoplasmatic membrane of living E. coli cells. From
these data we conclude that they reside in distinct and dynamic membrane domains,
which might be of functional importance.
References:
Pohl, T., Uhlmann, M., Kaufenstein, M. & Friedrich, T., Biochemistry 2007, 46,
10694-10702.
Lenn, T., Leake, M. C., Mullineaux C. W., Mol Microbiol. 2008,70,1397-407.
Mileykovskaya, E. & Dowhan, W., J. Bacteriol. 2000, 182, 1172-1175.
Johnson, A. S., van Horck, S. & Lewis P. J., Microbiology 2004, 150, 2815-2824.
9

Factors controlling stress perturbations in faulted reservoirs -
insights from numerical parameter studies
Fischer, K. & Henk, A.
Institut für Geowissenschaften – Geologie, Albert-Ludwigs-Universität Freiburg, Germany
karsten.fischer@geologie.uni-freiburg.de
Das tektonische Spannungsfeld beeinflusst die optimale Erschließung einer Lagerstätte in
vielfältiger Weise. So hängen u.a. Bohrlochstabilität, Orientierung von hydraulisch induzierten
Fracs und – insbesondere in geklüfteten Reservoiren – Permeabilitätsanisotropien von
den rezenten in situ Spannungen ab. Allerdings kann das regionale Spannungsfeld durch
Störungen und Lithologiewechsel lokal in Magnitude und Orientierung variieren. Insbesondere
bei störungskontrollierten Lagerstätten können solche Perturbationen dazu führen, dass
das Spannungsfeld innerhalb einzelner Störungsblöcke deutlich vom regionalen Trend abweicht.
Eine belastbare „pre-drilling“ – Prognose von tektonischen Spannungen zur Minderung
des Erschließungsrisikos und zur Optimierung der Bohrungen erfordert daher die Einbeziehung
der spezifischen Lagerstättengeometrie sowie der mechanischen Eigenschaften
der beteiligten Lithologien und Störungen. Als Prognosewerkzeug bieten sich geomechanische
Lagerstättenmodellierungen auf Basis der Finite Elemente (FE) Methode an.
Im Rahmen des DGMK Forschungsprojekts 721 wird das Potential von solchen geomechanischen
Lagerstättenmodellen für die Prognose von Spannungsfeldern und Kluftnetzwerken
am Beispiel des Erdgasfeldes Nord Hannover untersucht. In der ersten Projektphase wurden
im Rahmen von Parameterstudien die wesentlichen Kontrollfaktoren für Spannungsfeldperturbationen
an Störungen bearbeitet. Ausgehend von einem Standardmodell, das sich hinsichtlich
der idealisierten Geometrie eines Störungsblocks, der Materialeigenschaften und
Randbedingungen bereits an die Fallstudie Nord Hannover anlehnt, wurden systematisch die
Magnitude und die Orientierung der maximalen regionalen Hauptnormalspannung (σHmax),
die Reibungskoeffizienten der Störungen sowie die mechanischen Eigenschaften der Gesteine
variiert. Darüber hinaus wurden Modellszenarien mit mehreren Störungsblöcken bzw.
unterschiedlichen Größen der Störungsblöcke untersucht und zwei- und dreidimensionale
numerische Simulationen miteinander verglichen. Für alle Szenarien wurden die Ergebnisse
in Form von Isolinien- und Vektorplots für unterschiedliche Ausgabegrößen von Spannung
und Deformation (Magnituden und Orientierungen der Hauptnormalspannungen, von Mises
Vergleichsspannung, mittlere Spannnung, Differentialspannung, elastische und plastische
Deformationsanteile, „fault slip and dilation tendencies“) dokumentiert.
Die Parameterstudien zeigen, dass die Reibungskoeffizienten der Störungen, der Winkel
zwischen Störung und σHmax sowie die mechanischen Eigenschaften der aneinandergrenzenden
Gesteine den relativ größten Einfluss auf Spannungsfeldperturbationen haben. Während
das Standardmodell an den Störungen lediglich kleinräumige Veränderungen in der
Magnitude der Normalspannungen zeigt, haben z.B. deutlich herabgesetzte Reibungseigenschaften
ausgeprägte Veränderungen in Magnitude und Orientierung des Spannungsfeldes
zur Folge. Die Ergebnisse dieser Parameterstudien werden in der nächsten Projektphase zur
Validierung eines detaillierten Lagerstättenmodells von Nord Hannover genutzt, das die spezifische
Störungsgeometrie und mechanische Gesteinseigenschaften berücksichtigt.
10

Cloning, Expression, Purification and Crystallization of the First
Archaeal 4-Hydroxyphenylpyruvate Dioxygenase
Eduard Frick, 1 Thomas Spatzal, 2 Oliver Einsle, 2
Michael Müller, 1 and Wolfgang Hüttel 1
1 Institut für Pharmazeutische Wissenschaftten der Albert-Ludwigs-Universität Freiburg,
Lehrstuhl für Pharmazeutische und Medizinische Chemie, Albertstr. 25, D-79104 Freiburg,
Email: wolfgang.huettel@pharmazie.uni-freiburg.de
2 Institut für Organische und Biochemie der Albert-Ludwigs-Universität Freiburg,
Lehrstuhl für Biochemie, Albertstr. 21, D-79104 Freiburg,
Email: einsle@bio.chemie.uni-freiburg.de
4-Hydroxyphenylpyruvate dioxygenase (Hpd) catalyzes the conversion of
4-hydroxyphenylpyruvate (4-HPP) into 2,5-dihydroxyphenylacetate (homogentisate). This
reaction is part of the aerobic tyrosine catabolism in nearly all forms of life. Hpd is a valuable
target for agrichemical (development of herbicides) and therapeutical (treatment of hereditary
tyrosinaemia) research. [1]
Enzymes of extremophilic organisms offer a promising access to biocatalysts with increased
stability. We focussed on the Hpd from the acidophilic and mesothermophilic archaeon
Picrophilus torridus. [2] We managed to clone, express, purify, characterize and crystallize the
protein. The structure of P. torridus Hpd was determined at 2.3 Å resolution and will provide
further insights into the reasons for acid stability of certain proteins.
References:
[1] G. R. Moran, Arch. Biochem. Biophys. 2005, 433, 117-128.
[2] C. Schleper, et al., J. Bacteriol. 1995, 177, 7050-7059.
11

Tag der Forschung 2011
Laurdan labeled liposomes
– a new approach to quantify drug partitioning
Gesche Först, Rolf Schubert
Dept. of Pharmaceutical Technology and Biopharmacy, University of Freiburg
gesche.foerst.de@pharmazie.uni-freiburg.de
The knowledge of the partition behaviour of a drug between buffer and biological
membranes is important to predict its distribution and accumulation in the human
organism. Liposomes are common membrane models and ideal systems for the
characterization of drug-membrane interactions [1] .
The aim of this project is to establish an indirect screening method to investigate the
liposome/buffer partition coefficient of drugs using the easily available fluorescence
dye 6-Lauryl-2-dimethylaminonaphtalene (Laurdan) [2] .
After investigating the partitioning behaviour of bile salts as amphiphilic model drugs
in previous studies [3] the Laurdan method has also been applied to the ß-blockers
as additional substance class and the attained results are comparable to literature [4] .
Plemper v. Balen G, Marca Martinet C, Caron G, Bouchard G, Reist M, Carrupt PA, Fruttero R, Gasco A, Testa B.
Liposome/water lipophilicity: methods, information content, and pharmaceutical applications. Med Res Review. 2004; 24(3):
299-324. doi:10.1002/med.10063
[2] Paternostre M, Meyer O, Grabielle-Madelmont C, Lesieur S, Ghanam M, Ollivon M. Partition coefficient of a surfactant
between aggregates and solution: application to the micelle-vesicle transition of Egg-phosphatidylcholine and Octyl-�-Dglucopyranosid.
Biophys J 1995; 69: 2476-2488
[3] Theobald A, Thesis, Laurdan als Fluoreszenzsonde zur Quantifizierung der Wechselwirkung von Gallensalzen mit
Liposomenmembranen 2009; University of Freiburg
[4] Liu X., Hefesha H., Scriba G., Fahr A. Retention Behavior of Neutral and Positively and Negatively Charged Solutes on
an Immobilized-Artificial-Membrane Stationary Phase Helv. Chim. Acta 2008; 91: 1505 - 1512
12

Mechanistical studies of the Rhodium catalyzed
Hydroformylation with Self-Assembling Ligands
Urs Gellrich, Bernhard Breit*
Institut für Organische Chemie und Biochemie,
Albert-Ludwigs-Universität Freiburg, Albertstr. 21, 79104 Freiburg
Selectivity control in homogeneous metal complex catalysis is most frequently
achieved by crafting the microenvironment of the catalytically active metal center
upon binding of appropriate ligand architectures to the metal center. Among the
many ligands used, bidentate ligands occupy an important position. During the last
years our group was able to apply self-assembling ligands, interacting via hydrogen
bonds, [1] successfully in many catalytic reactions, for example hydroformylation,
hydrogenation and others. [2,3,4] In order to improve catalytic reactions and ligands it is
by all means necessary to completely understand the nature of the catalytically active
species. Hence, online monitoring of progressing catalytic transformations, supported
by quantum mechanical calculations can deliver valuable informations on the
intermediates involved in catalysis. We herein report on our first insights into
hydroformylation reactions with the 6-DPPon ligand using online
ReactIR TM -spectroscopy together with DFT calculations.
Figure 1: Direct observation of the formation of an intermediat of the catalytic cycle and the
corresponding DFT calculated structure (B3P86/def2-SVP).
[1] W. Seiche, B. Breit Angew. Chem., Int. Ed. 2005, 44, 1640.
[2] C. Waloch, J. Wieland, M. Keller, B. Breit, Angew. Chem. 2007, 119, 309, Angew. Chem. Int. Ed.
2007, 46, 3037.
[3] J. Wieland, B. Breit Nature Chemistry 2010, 2, 832.
[4] M. deGreef, B. Breit Angew. Chem., Int. Ed. 2009, 48, 559.
13

Streptomyces calvus – from bald to beautiful
Arne Gessner
Institut für pharmazeutische Biologie und Biotechnologie
Albert-Ludwigs-Universität Freiburg
arne.gessner@pharmazie.uni-freiburg.de
Streptomyces calvus is most prominently known for producing the fluorinated compound
nucleocidin. S. calvus is also a non-sporulating strain, an unusual property for Streptomycetes, which
is reflected in the Latin word for bald, ‘calvus’. Examination of a draft sequence of the genome of this
strain revealed a polyketide-biosynthesis gene cluster that has high sequence similarity to known
polyene-biosynthesis clusters like those for Nystatin or Pimaricin, which are valuable natural
products that are therapeutically used to treat fungal infections in humans and animals.
A leucine in the loading domain of a PKS in this biosynthesis cluster is encoded by a TTA-codon. Such
codons are extremely rare in the GC-rich genomes of Streptomycetes. The corresponding Leu-tRNA is
encoded by the bldA gene. Sequence alignment of the S. calvus bldA gene with bldA genes from
other Streptomyces species showed a point mutation that is predicted to encode a mis-folded and
most likely non-functional Leu-tRNA (TTA). So it was hypothetisized that complementation of S.
calvus with a functional copy of the bldA gene would not only restore sporulation but could also
activate the cryptic polyene cluster. The S. calvus bldA(+) strain did indeed regain the ability to
sporulate. Additionally, LC-DAD-MS analysis of an extract from a seven-day production culture
revealed a new peak that showed an UV-spectrum typical for a polyene. The corresponding polyene
‘annimycin’ was purified and the structure elucidated by 1 H-, 13 C-, HSQC, HMBC and ROESY NMR
spectroscopy. Disruption of PKS genes are currently underway to define the biosynthetic cluster for
annimycin.
14

UNRAVELING THE BIOSYNTHESIS OF KOTANIN AND ITS MONOMERIC PRECURSOR 7-
DEMETHYLSIDERIN IN ASPERGILLUS NIGER
C. Gil Girol, W. Hüttel, M. Müller
Institute for Pharmaceutical Sciences, University of Freiburg
Email: michael.mueller@pharmazie.uni-freiburg.de
A. niger shows an intermolecular stereo- and regioselective oxidative phenol coupling of the
coumarin derivative 7-demethylsiderin (2), forming orlandin (5). Final O-methylation leads to kotanin
(6) (Fig.1). 1 The biosynthesis of kotanin (6) in A. niger and its absolute configuration was elucidated
through feeding experiments and stereoselective synthesis. 1, 2 Up to now there is no evidence for an
alternative biosynthetic route to 6 by oxidative coupling of 3,7-dihydroxycoumarin (1) leading to the
bicoumarin 4 and subsequent O-methylation.
HO
HO
HO
CH 3
CH 3
CH 3
OH
13
C
O O
OH
13
C
O O
O 13C O
OH
HO
CH 3
O 13 CH 3
13
C
O O
H 3CO
1 2 3
?
?
HO
HO
CH 3
CH 3
O 13 CH 3
13
C
O O
O 13C O
OCH 3
H 3CO
H 3CO
4 5 6
Figure 1: Biosynthesis of kotanin (6) elucidated through feeding experiments and stereoselective synthesis.
CH 3
CH 3
CH 3
O 13 CH 3
13C O O
O 13 CH 3
13
C
O O
O 13C O
OCH 3
Here, we report the verification of this biosynthetical pathway on genomic level by a gene targeting
approach. In preliminary feeding experiments the polyketidic orign of kotanin was proven, an in silico
analysis of the A. niger genome 3 revealed putative PKS clusters coding for the monomeric precursor
2. Subsequent disruption of the genes led to the PKS responsible for kotanin (6) biosynthesis.
Furthermore disruption of an O-methyl transferase of the cluster provided evidence for absence of
an alternative route to 6 via 1 and 4.
1. Hüttel, W. & Müller, M. Regio- and stereoselective intermolecular oxidative phenol coupling
in kotanin biosynthesis by Aspergillus niger. Chembiochem 8, 521-529 (2007).
2. Hüttel, W., Nieger, M. & Müller, M. A short and efficient total synthesis of the naturally
occurring coumarins siderin, kotanin, isokotanin A and desertorin C. Synthesis, 1803-1808
(2003).
3. Pel, H.J. et al. Genome sequencing and analysis of the versatile cell factory Aspergillus niger
CBS 513.88. Nature Biotechnology 25, 221-231 (2007).
15
1, 2

GAP: A Genome Annotation Pipeline
Grüning BA, Erxleben A, Senger C, Flemming S, Günther S*
Pharmaceutical Bioinformatics, Institute of Pharmaceutical Sciences, University of
Freiburg, Germany
*e-mail: stefan.guenther[at]pharmazie.uni-freiburg.de
Introduction: Processing the exploding number of new sequence data requires efficient
implementation of several specialised tools on a powerful hardware infrastructure. Employed
methods include genome assembly, genome annotation, pathway reconstruction, data
visualisation, and pathway modelling. Galaxy is a workflow management system for data
processing ideally suited to the combination of various software tools. Furthermore, it ensures data
and process reproducibility in terms of repeatability and traceability. Accessibility via web interface
facilitates the integration of Galaxy into genome annotation projects.
Methods and Results: Based on Galaxy, we have developed the comprehensive Genome
Annotation Pipeline (GAP) focused on processing newly sequenced bacterial genomes. Currently,
the pipeline includes several publicly available and self-developed tools: Glimmer3 for the
prediction of ORFs, similarity searches in NCBI databases and UniProt, InterProScan for the
functional annotation of gene products, SignalP for the prediction of protein localisation, Aragorn
for tRNA and tmRNA detection within the genome and many more. The pipeline is implemented on
a high-performance computer and allows a very fast complete annotation of an average microbial
genome. GAP has been successfully applied for the annotation of Streptomyces TÜ6071. This is a
bacterium which has a highly-active isoprenoid biosynthesis and produces the industrial important
terpene Phenalinolactone which has anti-bacterial activity against several Gram-positive bacteria.
The annotated genome sequence of Streptomyces TÜ6071 is now available under the NCBI
accession number AFHJ00000000.
Future Prospects: Additional sequence analysis tools and and interfaces for complex data
visualisation will be implemented. GAP will soon be publicly available for genome annotation and
systems biology for the scientific community.
16

Strukturelle Charakterisierung von Br9 - -Anionen
Heike Haller, Mathias Ellwanger, Sebastian Riedel*
Institut für Anorganische und Analytische Chemie
Uni Freiburg
Seit Jahrzehnten ist eine Vielzahl von Polyiodidanionen bekannt und strukturell untersucht.
Im Bereich der niedrigeren Homologen kennt man deutlich weniger Beispiele. So sind bei den
Polybromiden mehrere Kristallstrukturen bekannt wie zum Beispiel Br10 2- , (Br - )2(Br4 2- ) und
ein unendliches 2D Polybromidnetzwerk. Anfang diesen Jahres wurde von Br20 2- als bisher
bromreichste bekannte Verbindung berichtet [1] . Jedoch ist im Bereich der
Polybromidmonoanionen lediglich Br3 - seit … als Kristallstruktur bekannt. Zu Br5 - , Br7 - und
Br9 - gibt es ausführliche ramanspektroskopische Untersuchungen [2] , eine genauere
kristallographische Untersuchung gelang jedoch bislang nicht.
Uns gelang erstmals NMe4Br9 sowie NEt4Br9 mittels Einkristallstrukturanalyse zu
charakterisieren.
[1] Wolff, M.; Meyer, J.; Feldmann, C.; Angewandte Chemie 2011,123, 5073-5077.
[2] Chen, X.; Rickard, M.; Hull Jr., J.; Zheng, C.; Leugers, A.; Simoncic, P.; Inorganic Chemistry 2010, 49,
8684-8689.
17

Gallium(I) Chemistry and the Synthesis of Highly Reactive Polyisobutylene
From Fundamentals to Applications
Higelin, A. and Lichtenthaler, M. R., Freiburg i. Br. / D
Prof. Dr. Ingo Krossing, Albert-Ludwigs-Universität Freiburg, Alberstraße 21,
79104 Freiburg i. Br. / D
In 2010 a simple route to univalent gallium salts of
weakly coordinating anions (WCAs) was developed
by SLATTERY et al. [1] Herein the gallium(I)-cations are
solely coordinated by aromatic solvent molecules (e.g.
toluene or fluorobenzene) in a bent sandwich fashion.
As only one Ga-F contact is shorter than the sum of
the van der Waals radii (3.34 Å) the interaction
between the gallium(I)-cations and the WCAs is very
weak (Figure 1).These novel salts are therefore ideal
starting materials for further gallium(I) chemistry.
The solvent molecules can be replaced by a variety of
σ-donor ligands, like phosphanes and ethers, leading
to an entirely new class of coordination compounds
Figure 1: Molecular Structure of
[Ga(C 6H 5Me) 2] + [Al(OC(C(CF 3) 3) 4] – .
which were inaccessible with previously known Ga(I) starting materials. The study of these
exciting new compounds and their exotic bonding situations, in conjunction with quantum
chemical investigations, gives fundamental insights into the nature of the chemical bond. On the
applied side, the variation of the ligands offers an elegant method of tuning the reactivity of the
gallium(I)-cations. The latter namely are a promising species for a variety of applications, e.g.
initiating or catalyzing the synthesis of highly reactive Polyisobutylene (HR-PIB).
Depending on its molecular weight Polyisobutylene (PIB) has been used in many versatile
applications. Thus high molecular weight PIB has gradually substituted natural chicle as the
main chewing gum base. The application of low molecular weight PIB however relies on the
PIB being highly reactive. PIB is classified as highly reactive as soon as the content of terminal
double bonds exceeds 60%. Being sterically exposed, the latter can easily be functionalized.
Therefore HR-PIB represents an essential intermediate in manufacturing additives for lubricants
and fuels. [2] Up to this day most syntheses of HR-PIB show a lack in at least one point. May it
be low reaction temperatures down to –100 °C, dichloromethane as solvent or relatively high
concentrations of the initiating or catalyzing species – all these factors sum up to a negative
[3], [4]
economic and environmental balance.
Recently a number of reactions showed the superior quality of the univalent gallium salts
initiating or catalyzing the polymerization of isobutylene. Thus the synthesis of HR-PIB can be
carried out at relatively high reaction temperatures up to 0 °C, in a non carcinogenic and non
water-hazardous solvent, i.e. toluene, as well as using relatively low concentrations of the
initiating or catalyzing species, down to 0.007 mol%. The experimental results were backed by
quantum-chemical calculations giving a first hint on a coordinative polymerization mechanism.
[1] J. M. Slattery, A. Higelin, T. Bayer, I. Krossing, Angew. Chem. Int. Ed., 2010, 49, 3228.
[2] M. Vierle, Y. Zhang, E. Herdtweck, M. Bohnenpoll, O. Nuyken, F. E. Kühn, Angew. Chem. Int Ed. 2003, 42, 1307.
[3] A. Guerrero, K. Kulbaba, M. Bochmann, Macromol. Chem. and Phys. 2008, 209, 1714.
[4] Y. Li, H. Y. Yeong, E. Herdtweck, B. Voit, F. E. Kühn, Eur. J. Inorg. Chem. 2010, 4587.
18

Characterization of airborne dust particles in the coal mining area of Cam Pha,
northern Viet Nam
T.B. HOÀNG-HÒA 1* , R. GIERÉ 1 , V. DIETZE 2 , U. KAMINSKI 2 AND P. STILLE 3
1 Albert-Ludwigs-Universität, D-79104 Freiburg, Germany (*correspondence: hoahtb@gmail.com)
2 German Meteorological Service, Research Center Human Biometeorology, Stefan-Meier-Str. 4, D-79104, Freiburg, Germany
3 École et Observatoire des Sciences de la Terre, Université de Strasbourg, F-67084 Strasbourg, France
Cam Pha, located in Quang Ninh province, is one of the largest coal mining areas in Viet Nam
(reserves ~10 Gt). This study focuses on the mineralogical and chemical characterization of
airborne dust. Exposure to dust is a major challenge for the population living near the mines,
especially the open-pit mines. Additional major dust sources comprise mine dumps containing the
overburden, a coal-processing facility, a coal-shipping harbour, mining traffic, and various
industries, including coal-fired power stations.
Coarse particles (dp >2.5 µm) have been collected with the passive sampler device Sigma-2 on
transparent adhesive collection plates for subsequent single-particle analysis by automated optical
microscopy according to VDI guideline 2119 [1]. Select specimens, sampled during different
meteorological conditions, were investigated further by SEM-EDX single-particle analysis and by
determining their bulk chemical and isotopic composition (ICP-MS and MC-ICP-MS, respectively).
Wind directions indicate that the particles are mainly derived from the open-pit mines, consistent
with bulk chemical and isotopic data ( 87 Sr/ 86 Sr = 0.7278 – 0.7427; εNd = -14.6 – -14.9; 206 Pb/ 207 Pb =
1.1789 – 1.1884), which suggest that the dust contains mostly natural materials (coal, silicate
minerals from sedimentary rocks). Relative to the coal, the bulk airborne dust is enriched in Na,
Mg, K, and Ca, but depleted in most other components. Element-ratio plots reveal some systematic
differences between the dust samples and specimens of coal and overburden, pointing to an
additional, yet unknown particle source. To verify the hypothesis of an additional dust source and to
provide quantitative data on the mineralogical composition of the dust samples, we are currently
optimizing the automated SEM-EDX single-particle analysis technique.
[1] VDI (1997): VDI guideline 2119, part 4.
19

Towards the Total Synthesis of β-Lipomycin:
A Highly Convergent and Adaptable Double Stille-Coupling
Strategy for the Synthesis of 3-(Polyenoyl)tetramic Acids
Max Hofferberth and Reinhard Brückner*
Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität, Albertstraße 21,
79104 Freiburg, Germany; E-mail: max_hofferberth@web.de
3-(�-Hydroxymethylidene)pyrrolidine-2,4-diones constitute the major tautomer of structures
commonly referred to as „acyltetramic acids“. Compounds in which „acyl“ equals „poly-
enoyl“ define a class of natural products, which continues to evoke interest both in the
synthetic [1] and biosynthetic community [2] .
We synthesized a prototypical (polyenoyl)tetramic acid, namely β-lipomycin (9), [3] � or pos-
sibly a stereoisomer thereof, since the 3 D structure of 9 still awaits elucidation. Employing a
novel approach, which is variable and convergent the naturally occurring stereoisomer plus
chain-extended or chain-altered derivatives thereof should become readily accessible.
[1] R. Schobert, A. Schlenk, Bioorg. Med. Chem. 2008, 16, 4203-4221 and literature cited therein.
[2] E. g.: C. Bihlmaier, E. Welle, C. Hofmann, K. Welzel, A. Vente, E. Breitling, M. Müller, S. Glaser, A.
Bechthold, Antimicrob. Agents Chemother. 2006, 50, 2113-2121.
[3] B. Kunze, K. Schabacher, H. Zähner, A. Zeeck, Arch. Mikobiol. 1972, 86, 147-174.
[4] First synthesis and applications of 6: A. Sorg, R. Brückner, Angew. Chem. 2004, 116, 4623-4626; Angew.
Chem. Int. Ed. 2004, 43, 4523-4526; J. Burghart, R. Brückner, Eur. J. Org. Chem. 2011, 150-165; J.
Burghart, A. Sorg, R. Brückner, Chem. Eur. J. 2011, 17, 6469-6483.
20

Chorismatases: Characterizing a new group of enzymes
F. Hubrich, J. Andexer
Institut für Pharmazeutische Wissenschaften, Pharmazeutische und Medizinische Chemie, Albertstr.
25, 79104 Freiburg
Chorismatases are enzymes catalyzing the hydrolysis of chorismic acid 1 to pyruvat and a cyclic
carboxylic acid. The level of saturation of the carboxylic acid depends on the type of enzyme
catalyzing this hydrolysis. Enzymes of the FkbO-type produce 1,5-diene-3,4-dihydroxycyclohexanoic
acid (DHCHC) 3, enzymes of the Hyg5-type produce the aromatic 3-hydroxybenzoic acid (3HBA) 2. 1
These products are precursor molecules for the synthesis of natural products like ascomycin 5 ,
rapamycin 2 , brasilicardin 3 and xanthomonadin 4 .
FkbO and Hyg5 are kinetically characterized for chorismate, but both the structure and the reaction
mechanism of these enzymes are still unknown. Structural modeling will be difficult, because there is
no sequence similarity between chorismatases and other chorismate using enzymes.
Phylogenetic analysis 1 and sequence alignments (BLAST) show additional genes in different
organisms (Streptomyces and Xanthomonas), that are probably encoding for chorismatases. To show
that these genes products are chorismatases, the target genes are introduced into E. coli expression
vectors and strains. The already purified protein FkbO and Hyg5 are used in crystallization
experiments to get a crystal structure of a chorismatase.
21

Lawsone and Tetrahydroxynaphthalene Reductase
Syed Masood Husain, Michael Schätzle, Michael Richter, Michael Müller
Institut für Pharmazeutische Wissenschaften der Albert-Ludwigs-Universität Freiburg,
Albertstr. 25, D-79104 Freiburg im Breisgau, Email: michael.mueller@pharmazie.uni-freiburg.de
Lawsone (2-hydroxy-1,4-naphthoquinone), is a red-orange dye present in the leaves of
the henna plant (Lawsonia inermis). Humans have used henna extracts containing
lawsone as hair and skin pigments for more than 5000 years. We have used this natural
product as a substrate, which on reduction catalyzed by the enzyme called
tetrahydroxynaphthalene reductase (T4HNR) from Magnaprothe grisea, gave cis-keto-
diol with high yield (95%) and high ee (99%).
HO
O
O
Lawsone
T 4HNR
NADPH
HO
O
OH
cis-keto-diol
Lawsone was also used extensively to study the mechanism of T4HNR-catalyzed
reduction that has led to the formation of a cis-keto-diol.
22

Stereoselectivity of the Ketoreductase Tyl-KR1 from
Streptomyces fradiae — A VCD spectroscopy analysis
Matthias Häckh, Michael Müller, and Steffen Lüdeke
Institut für Pharmazeutische Wissenschaften der Albert-Ludwigs-Universität Freiburg,
Albertstr. 25, D-79104 Freiburg, Germany
Enzyme-catalysed reactions are best-known for their high stereoselectivity. Bio-
catalysis benefits from these outstanding properties making chiral building blocks
accessible that can be used for drug syntheses. [1] Oxidoreductases play an important
role in polyketide chemistry, especially those with rarely observed “anti-Prelog type”
selectivity, since oxidoreductase-catalysed reduction of ketones generally leads to
products with “Prelog type” configuration. [2] The stereoselectivity of many enzymes is
substrate-specific and might be decreased or even converted for non-physiological
substrates. Exact determination of the absolute configuration of different products
formed out of different substrates is therefore very important for the useful application
of enzymes in biocatalysis. A switch between “anti-Prelog type” and “Prelog type”selectivity
seems to be possible.
The ketoreductase Tyl-KR1 from Streptomyces fradiae is known to reduce racemic 2methyl-3-oxopentanoic
acid N-acetylcysteamine thioester (1) enantioselectively to
anti-(2R,3R)-3-hydroxy-2-methylpentanoic acid N-acetylcysteamine thioester (2). [3]
This “anti-Prelog type” anti-configuration is rarely found in natural products.
O O
CH 3
S
H
N
TYL KR 1
OH
R
O
S
1
O
CH3 2
anti-R,R
R
"anti-Prelog type"-reduction
Figure 1: “Anti-Prelog type”-reduction catalysed by Tyl-KR1.
We used VCD spectroscopy and quantum chemical calculations to determine the
absolute configurations of four different polyketide analogues reduced by Tyl-KR1.
References
[1] M. Müller, Angew. Chem. Int. Ed. 2005, 44, 362.
[2] K. Faber, Biotransformations in Organic Chemistry, 5th ed., Springer, 2004.
[3] A. P. Siskos, A. Baerga-Ortiz, S. Bali, V. Stein, H. Mamdani, D. Spiteller, B. Popovic,
J. B. Spencer, J. Staunton, K. J. Weissman, P. F. Leadlay, Chem. Biol. 2005, 12,
1145.
23
H
N
O

Engineering an aryl-C-glycoside biocatalyst
J. Härle 1 , M. Weber 1 , B. Lauinger 1 , S. Günther 2 , B. Kammerer 3 , A. Luzhetskyy 1 , A. Bechthold 1 *
(1) Department of Pharmaceutical Biology und Biotechnology
(2) Department of Pharmaceutical Bioinformatics, Institute of Pharmaceutical Sciences, Albert-Ludwigs-
Universität Freiburg, 79104 Freiburg, Germany.
(3) Zentrum für Biosystemanalytik, Albert-Ludwigs-Universität Freiburg, 79104 Freiburg, Germany.
*andreas.bechthold@pharmazie.uni-freiburg.de
Before a natural product potentially becomes a therapeutic agent properties influencing the
pharmacological or pharmacokinetic qualities might have to be improved. As sugar substituents play an
essentiell role in respect to bioactivity and solubility, they represent a valuable address for modification
(Harle and Bechthold, 2009).Since the sugar moieties are primarily O-linked to the natural product core the
acid sensitivity strictly limits a principal therapeutical application. A practical alternative is to improve the
molecule utilizability by replacing the labile O-aryl bond with an enzymatical and chemical stable Cglycosidic
bond. The resulting structural difference might not influence the bioactivity but raises
enormously the rigidity. The observed flexibility of UrdGT2 catalyzing C- and O-glycosidic bonds (Durr et al.,
2004) brought up the idea to engineer LanGT2, a strictly O-glycosyltransferase involved in the biosynthesis
of the anticancer agent landomycin A (Luzhetskyy et al., 2005),towards a C-glycosylating enzyme.
By integrating computational assistance with multiple target amino acid replacement experiments, we
interconverted the naturally O-glycosylating enzyme LanGT2 towards a glycosyltransferase evolved to
catalyzes the sugar attachment by a C-glycosidic bond. Elucidated by alanin substitutions and visulized by in
silico protein modeling we illustrate the assumed catalytic mechanism determining the C- and Oglycosylation
respectively. The achievement in overcoming natural limitations by protein engineering might
document a general strategy in gaining further chemical diversity in the field of combinatorial biosynthesis.
References:
Durr,C., Hoffmeister,D., Wohlert,S.E., Ichinose,K., Weber,M., Von Mulert,U., Thorson,J.S., and Bechthold,A.
(2004). The glycosyltransferase UrdGT2 catalyzes both C- and O-glycosidic sugar transfers. Angew. Chem.
Int. Ed Engl. 43, 2962-2965.
Harle,J. and Bechthold,A. (2009). Chapter 12. The power of glycosyltransferases to generate bioactive
natural compounds. Methods Enzymol. 458, 309-333.
Luzhetskyy,A., Taguchi,T., Fedoryshyn,M., Durr,C., Wohlert,S.E., Novikov,V., and Bechthold,A. (2005).
LanGT2 Catalyzes the First Glycosylation Step during landomycin A biosynthesis. Chembiochem. 6, 1406-
1410.
24

Master Thesis: The Influence of Mechanical Defects on the Lattice of silicon
T. Jauss 1 , J. Wittge 1 , A.N. Danilewsky 1 , A. Cröll 1 , J. Garagorri 2 , R. M. Elizalde 2 , D. Allen 3 , P. McNally 3
1
Universität Freiburg, Kristallographie, Geowissenschaftliches Institut, Freiburg, Germany
2
Centro de Estudios e Investigaciones Tecnicas de Gipuzkoa, CEIT and Tecnun (University of Navarra), San
Sebastian, Spain
3
Dublin City University, The RINCE Institute, Dublin, Ireland
Correspondence e-mail: thomas.jauss@jupiter.uni-freiburg.de
The aim of this Master was thesis to examine the impact of mechanical damage to the silicon lattice. For this
reason controlled damage was applied to silicon wafers with a boron doped layer by nanoindentation [Gar10]. A
diamond Vickers tip on an Agilent Nanoindenter II has been used for indentation and the resulting damage has
been characterized by differential interference contrast microscopy (NIC) and high resolution X-ray diffraction
(HRXRD). Damage by loads of 1N – 5N is compared.
Rocking curves (RC) and reciprocal space maps (RSM) were recorded and analyzed with respect to full width at
half maximum and lattice plane spacing by the method of Bond. The volume of the disturbed crystal material
around the indents is very small compared to the undisturbed volume. Therefore a line of seven indents with 1N
indentation load is oriented in a way that it lies parallel to the line focus of the X-ray beam in order to increase
the signal intensity of the disturbed crystal. The RSM of the undisturbed crystal volume is regarded as a
reference, the sharp RC are confirmed by the sharp confined peaks in the RSM. Only artefact streaks of the
monochromator and the analyzer are also clearly visible, as well as the dynamical streak of the specimen (Fig.
1a). Despite the low count rate, a clear anisotropic intensity distribution around the layer related peak is visible at
the damaged area (Fig. 1b).
Fig. 1: Reciprocal space maps of the 004 reflection. Two peaks produced by the lattice mismatch between the
silicon substrate and the boron doped layer can be observed in both cases.
a) Undisturbed area: sharp confined peaks of the silicon substrate and the B-doped layer, as well as the apparatus
induced artefact streaks and the dynamical specimen streak.
b) Disturbed area: an anisotropic broadening of the peak bases is clearly visible, but mainly concentrated around
the layer related peak.
[Gar10] Garagorri, J., Gorostegui-Colinas, E., Elizalde, M.R., Allen, D., McNally, P. (2010) Nanoindentation
induced silicon fracture and 3D modelling, Anales de Mecánica de la Fractura, 27, pages 559-564.
25

Biomonitoring of atmospheric particles on cypress leaves
- sampled in the vicinity of a nuclear plant-
R. KALTENMEIER 1* , R. GIERÉ 1 AND L. POURCELOT²
1
Albert-Ludwigs-Universität, D-79104 Freiburg, Germany
(*correspondence: giere@uni-freiburg.de)
²Institut de Radioprotection et de Sûreté Nucléaire, Cadarache,
F-13115 St Paul lez Durance, France
Environmental pollution can be assessed and evaluated by plants as biological monitors. Their use
can be selected depending on the investigation objective, by their ability to reflect past and/ or
current environmental conditions as well as temporal changes (Wolterbeek 2002). Coniferous trees
belong to the best passive bioindicators for air, soil and water pollution (Saito et al. 2004). For
cypress leaves (Chamaecyparis nootkatensis), sampled in the vicinity of the Malvesi unranium
processing facility, SW France, increased activity of actinides and some of their decay products has
been reported (Figure 1). Ejection via smokestacks and artificial ponds inside the facility are the
source of these enhanced activities. Air dried leaf samples were examined systematically by scanning
electron microscopy (SEM) in secondary and backscattered electron mode in combination with
energydispersive X-ray spectrometry (EDX), in order to characterize particulate matter deposited on
leaf surfaces and to verify the existence of radioactive particles. For the interpretation of qualitative
EDX spectra of individual particles trapped on the leaf surfaces, signals resulting from C or AU coating
as well as the background spectra of the cypress leaves themselves were taken into account (Figure
2). Particle sizes ranging from

Carbamates as a new prodrug concept for HDACs
Keller, K. [a] , Stolfa, D.A., [a,d] Schlimme, S., [b] Hauser, A-T., [a] Carafa, V., [c] Heinke, R.,
[b] Kannan, S., [b] Cellamare, S., [d] Carotti, A., [d] Altucci, L., [c] Jung, M., * [a] and Sippl,
W., * [b]
[a] Albert-Ludwigs-Universität Freiburg, Albertstr. 25, 79104 Freiburg
[b] Martin-Luther-Universität, Wolfgang-Langenbeck-Str. 4, 06120 Halle/Saale
[c] Seconda Università di Napoli, Vico L. De Crecchio 7, 80138 Napoli
[d] Università degli Studi “Aldo Moro”, Via Orabona 4, 7012 Bari
Histone acetyltransferases (HATs) and histone deacetylases (HDACs) are
responsible for maintaining the acetylation equilibrium. HDACs are amidohydrolases
that cleave the acetamide bond by a zinc-dependent mechanism. Inhibitors of
HDACs have shown great promise in the development as new anticancer agents.
Hence, a prodrug principle for HDAC inhibitors is a valuable concept to optimize
cellular activity.
Our experimental results validate the general applicability of the carbamate group as
a prodrug for hydroxamates, as compound 2 is inducing cellular hyperacetylation in a
similar range of concentration as compared to the parent hydroxamate 1. These
results demonstrate that the introduction of a carbamate group is a valuable concept
for the design of new HDAC inhibitors with improved pharmacokinetic properties.
Introducing a carbamate group on the hydroxamic acid function of SAHA
(suberoylanilide hydroxamic acid), a drug already approved as a HDAC inhibitor, led
to further prodrugs, such as compound 3. We synthesized further carbamates from
SAHA with different moieties to further investigate stability, cellular activity and to
study the impact of different substitution patterns by considering pharmacokinetic
aspects like solubility.
Literatur:
S. Schlimme et al., ChemMedChem 2011, 6 (4)
27
31

STABILIZATION OF DIALKYL METAL COMPOUNDS OF GROUP 13 (R2E)2B12Cl12
(E = Al, Ga, In) USING THE WEAKLY COORDINATING DIANION [B12Cl12] 2-
Mathias Keßler, Ardiana Zogaj, Carsten Knapp
Institut für Anorganische und Analytische Chemie Albert-Ludwigs-Universität Freiburg,
79104 Freiburg i. Br., Germany; mathias.kessler@ac.uni-freiburg.de
The perchlorinated closo-dodecaborate [B12Cl12] 2- is a chemically very robust weakly
coordinating dianion, for which improved synthesis have been published recently.[1] It
was used for the syntheses of trialkyl silylium compounds (R3Si)B12Cl12 and the
methylating agent Me2B12Cl12 which indicate its potential to stabilize electrophilic
cations.[2]
The stabilization of ion-like dialkyl metal compounds of group 13 [R2E] + (E = Al, Ga,
In) is a challenging task. Bochmann et al. showed that the
tetrakis(pentafluorophenyl)borate [B(C6F5)4] – anion is not suitable as counterion
because degradation occurs during the reaction.[3] In 2002 Reed et al. succeeded in
the characterization of (Et2Al)(HCB11H5X6) (X = Cl, Br) using partly halogenated
carborane anions.[4]
The reactions of the trityl salt [CPh3]2[B12Cl12] with trialkyl metal compounds R3E of
group 13 (E = Al, In) and Et3Ga·(OEt2) leading to (R2E)2B12Cl12 (E = Al, In; R = Me,
Et) and [Et2Ga(OEt2)2]2[B12Cl12] will be discussed. Characterization of the compounds
using NMR- and Raman spectroscopy and X-Ray diffraction will be presented.[5]
Fig. 1 Part of the crystal structure of (Me2Al)2B12Cl12.
References:
[1] a) Knapp, C.; Guttsche, K.; Geis, V.; Scherer, H.; Uzun, R. Dalton Trans. 2009, 2687-2694. b) Gu,
W.; Ozerov, O. V. Inorg.Chem. 2011, 50, 2726-2728.
[2] a) Kessler, M.; Knapp, C.; Sagawe, V.; Scherer, H.; Uzun, R. Inorg. Chem. 2010, 49, 5223-5230. b)
Bolli, C.; Derendorf, J.; Keßler, M.; Knapp, C.; Scherer, H.; Schulz, C.; Warneke, J. Angew. Chem.
2010, 122, 3616-3619; Angew. Chem. Int. Ed. 2010, 49, 3536-3538.
[3] Bochmann, M.; Sarsfield, M. J. Organometallics 1998, 17, 5908-5912.
[4] Kim, K. C.; Reed, C. A.; Long, G. S.; Sen, A. J. Am. Chem. Soc. 2002, 124, 7662-7663.
[5] Kessler, M.; Knapp, C.; Zogaj, A. Organometallics 2011, accepted.
28

Untersuchung des Einflusses von Nanofüllstoffen auf die Trocknung und
die Eigenschaften von Polymerfilmen mittels forcierter Rayleighstreuung
Kiessling, Andy, Andy.Kiessling@fmf.uni-freiburg.de
Bartsch, Eckhard, Eckhard.Bartsch@physchem.uni-freiburg.de
Beschichtungen, welche auf der Verwendung von leicht flüchtigen organischen Substanzen
basieren, tragen u.a. zum bodennahen Ozon bei. Dieses verursacht eine Reizung des
respiratorischen Systems, welches bestehende Atemwegserkrankungen wie Asthma
verschlimmert. Im Gegensatz dazu entstehen bei der Herstellung von Beschichtungen aus
wässrigen Polymerdispersionen keine schädlichen Emissionen.
Trotz ihrer breiten Anwendung ist die Filmbildung aus wässrigen Polymerdispersionen
mechanistisch nicht in allen Details verstanden. Einer der wichtigsten Schritte dieser
Filmbildung ist die Teilchenkoaleszenz. Dabei lösen sich durch Interdiffusion der
Polymerketten die Teilchengrenzen auf und ein mechanisch stabiler Film entsteht. Dieser
Prozess wird in unserer Gruppe mit der Methode der forcierten Rayleighstreuung (FRS) auf
molekularer Ebene verfolgt. Hierbei wird die Diffusion eines Tracers über verschiedene
Weglängen beobachtet, indem durch die Überlagerung zweier Laserstrahlen ein
holographisches Gitter eingebleicht wird. Dessen Zerfall durch Farbstoffdiffusion wird über
das zeitliche Abklingen eines optischen Braggstreusignals detektiert. Aus der Interpretation
und Modellierung des Signals gewinnt man Informationen über die Mobilität des Tracers in
den verschiedenen Filmkompartimenten, welche durch Diffusionslängen und Diffusionskoeffizienten
charakterisiert wird.
Aktuell wird der Einfluss innerer und äußerer Partikel-Grenzflächen anhand von Kern-Schale-
Systemen untersucht. [1] Diese Untersuchungen sollen Aufschluss über den molekularen
Ursprung der makroskopisch beobachteten besonderen Eigenschaften wie Kratzfestigkeit
geben, die bei Beschichtungen aus Materialien komplementärer Eigenschaften auftreten.
Dafür werden die Schalendicke, die Kerngröße, die Art des Kerns und der Schale variiert.
Neben der Natur des Systems haben die Filmbildungsbedingungen (Temperatur,
Tensidkonzentration, Trocknungsgeschwindigkeit) enormen Einfluss auf die Eigenschaften
der gebildeten Beschichtung. Daher sind die Filmbildungsbedingungen ein weiterer wichtiger
Teil der Untersuchungen.
Links: Stadien der Filmbildung einer Latex-Dispersion, Mitte: elektronenmikroskopische Aufnahme der Teilchenkoaleszenz,
[2] Rechts: schematische Darstellung des FRS-Experiments.
[1] K. Suresh, E. Bartsch, J. Polym. Sci. B. 2007, 45, 2823-2834.
[2] Kolloide – Vorstoß in die Nanowelt, Topics in Chemistry, BASF AG, Ludwigshafen.
29

In vivo conversion of trans-3-hydroxy-L-proline by heterologously
overexpressed microbial trans-4-prolinehydroxylase
Christian Klein, Michael Müller and Wolfgang Hüttel
Lehrstuhl für Pharmazeutische und Medizinische Chemie, Albert-Ludwigs-Universität
Freiburg, Germany
christian.klein@pharmazie.uni-freiburg.de
Heterologously overexpressed proline hydroxylases provide the possibilities of enzymatic
conversions of L-proline for the regio- and stereospecific generation of hydroxyprolines,
useful chiral synthons for chemical synthesis of pharmaceuticals. [1,2] It was also found that
proline hydroxylases are not strictly substrate specific but accept a set of proline
congeners. [1,3]
Among several so far undiscovered reactions, proline-4-hydroxylase showed activity with
trans-3-hydroxy-L-proline resulting in the trans-2,3-cis-3,4-dihydroxyproline isomer
exclusively. Since in vitro conversion with the enzyme suffers from several drawbacks, an in vivo
approach was established. Product concentrations of up to 4 mM were obtained in the
fermentation broth of simple shake flask cultures and the product was isolated via ionexchange
chromatography in >90% yield.
References:
[1] C. Klein, W. Hüttel, Adv. Synth. Catal. 2011, 353, 1375-1383.
[2] P. Remuzon, Tetrahedron 1996, 52, 13803-13835.
[3] T. Shibasaki, W. Sakuri, A. Hasegawa, Y. Uosaki, H. Mori, M. Yoshida, A. Ozaki,
Tetrahedron Lett. 1999, 40, 5227-5230.
30

Optical excitations and initial charge transfer phenomena in
biopolymers and solar cell model compounds: a theoretical
approach
S. Kruse, S. Krapf, B. Lampe, T. Koslowski
Institut für Physikalische Chemie, Universität Freiburg, Albertstrasse 23a, D-79104 Freiburg
im Breisgau, Germany
We approach the problem of optical excitations in molecular aggregates in complex
biochemical and man-made environments from a computational, all-atom perspective. The
system is divided into a pi orbital part described by a Pariser-Parr-Pople model, which is
coupled to the charge degrees of freedom of a classical force field. Strategies for a highaccuracy
reparameterization and an efficient computational solution are presented.
For gamma-D-crystallin, a band edge consisting of charge-transfer states emerges for a
coupled molecular aggregate compared to the uncoupled residues. The energies of some
charge-transfer states strongly depend on the dielectric properties of the model, giving a first
insight into the potential temporal evolution of these excitations. Biochemical implications
and extensions of the model to solar cell model compounds are discussed.
31

Stabilized lipid disks
Lena Koehler 1 , Michael Müller 2 , Katarina Edwards 1,3 and Ulrich Massing 4
1 FRIAS, School of Soft Matter Research, Freiburg, Germany
2 Institute of Pharmaceutical Sciences, Albert-Ludwigs-Universität Freiburg, Germany
3 Physical and Analytical Chemistry, University Uppsala, Sweden
4 Tumor Biology Center, Freiburg, Germany
Background:
A novel type of membrane disks with promising characteristics have been found in lipid
mixtures containing polyethylene glycol (PEG)-lipids 1,2 . The lipid disks show planar and
circular shape. The PEG-lipids favor the rim of the disks 3 and offer steric protection against
fusion and self-closure. Lipid disks can be produced using the same techniques as for the
preparation of liposomes; such techniques like sonication or several freeze/thaw/vortexcycles
followed by extrusion give a very homogenous size distribution.
Stabilized lipid disks:
In order to realize the full potential of the disks increasing the robustness of the disks is
desirable. Therefore the formation of a stabilizing shell is envisioned. To meet this goal
modified PEG-lipids bearing a T-Linker between the lipid- and the PEG-moiety should be
synthesized. These modified PEG-lipids should be employed in a cross-linking reaction
between each other on the rim of the disks to enhance the cohesion and therefore the
stability of the discs.
References:
1. K. Edwards, M. Johnsson, G. Karlsson, M. Silvander, Biophys J 73 (1997), 258-266
2. M. Johnsson, K. Edwards, Biophys J 85 (2003) 3839–3847
3. P. Wessman, A. Rennie, K. Edwards, BBA Biomembranes 1778 (2008), 2210-2216A.
32

Implications of magnetite-mediated ROS formation on genotoxic
effects and signalling pathways in A549 cells
Mathias Könczöl 1 , Richard Gminski 1 , Sandra Ebeling 2 , Ella Goldenberg 3 , Reto Gieré 3 , Bernard
Grobéty 4 , Barbara Rothen-Rutishauser 5 , Irmgard Merfort 2 , Volker Mersch-Sundermann 1
1 Institut für Umweltmedizin und Krankenhaushygiene, Universitätsklinikum Freiburg
2 Lehrstuhl für Pharmazeutische Biologie und Biotechnologie, Universität Freiburg
3 Institut für Geowissenschaften, Universität Freiburg
4 Institut für Geowissenschaften, Universität Fribourg, Schweiz
5 Institut für Anatomie, Universität Berne, Schweiz
Magnetite, a ferromagnetic iron oxide, has been characterized as a major component
of particulate matter sampled at underground stations, along railway lines and at
welding workplaces. The abundance of magnetite in these samples and the lack of
consistent data demand more systematic investigation, specifically into the genotoxic
potential of magnetite nano- and micrometer particles and their underlying
mechanisms. In the present study, we studied magnetite for its toxic potential in
cultured human lung cells (A549). Cell viability (WST-1, NR assay), oxidative stress
(DCFH-DA, lipidperoxidation (LPO)) and genotoxicity (comet assay; CB-MNvit test)
were determined as biological endpoints of particle exposure. Additionally, effects on
NF-κB and JNK activation were evaluated by EMSA and western blots.
TEM images showed incorporation of magnetite bulk and nanoparticles into A549
cells. No cytotoxic effects were observed after exposure of the cells to the various
samples. We observed concentration-dependent ROS formation and as a
consecutive reaction high amounts of LPO for all samples. Preincuation with the ROS
scavengers butylated hydroxyanisole (BHA) and apocynin, known to inhibit NADPH-
oxidase (NOX) and to increase GSH-levels, lead to decreased ROS production. We
detected size and concentration-dependent induction of micronuclei (MN), as well as
DNA-damaging effects. These genotoxic effects could be reduced by the ROS
scavengers N-acetylcysteine and BHA. Moreover, an activation of JNK without
increased NF-κB-binding activity was observed. However, activation of JNK seems to
be not ROS dependent. Our experiments suggest that after endocytosis of magnetite
particles NOX-activation leads to ROS formation and consecutive LPO. These effects
may play an important role in the genotoxicity of magnetite in A549 cells.
33

Gemischte Thiooxowolframate A2[W VI SxO4−x]:
A. Lehner, M. Braitsch, Z. Deng, C. Röhr
Institut f. Anorg. u. Analyt. Chemie, Universität Freiburg
anna@limonite.chemie.uni-freiburg.de
Gemischte Thiooxoorthowolframate(VI) [WSxO4−x] 2− wurden erstmals im 19. Jahrhundert
dargestellt [1,2]. Die spektroskopischen Eigenschaften dieser Ionen wurden intensiv
untersucht [3], während kaum detaillierte Studien der Kristallstrukturen ihrer Salze vorliegen
[4]. Wir präsentieren hier die erste systematische Untersuchung der Kristallchemie
von Alkalimetall- und Ammoniumthiooxowolframaten anhand von Röntgendiffraktometrie
und Bandstrukturrechnungen. Die Strukturchemie und Bindungssituation wird dabei im
Vergleich mit den Ergebnissen unserer Untersuchung analoger Thiooxomolybdate [5,6] vorgestellt.
Die Synthesen der Salze A2[WSxO4−x] (A = NH4: x = 2; A = K, Rb: x = 1,2,3; A = Cs:
x = 2,3) erfolgte durch Einleiten von H2S-Gas in alkalische Lösungen der Oxowolframate.
Aus den gelben Thiooxowolframatlösungen ließen sich die Salze phasenrein und einkristallin
erhalten. Die Verbindungen weisen weit mehr polymorphe Formen auf als bisher erwartet
wurde: zusätzlich zu drei bekannten A2BX4-Typen wurden vier neue Strukturtypen erhalten.
Die Monothiowolframate von K + und Rb + kristallisieren als Hydrate. Während alle Verbindungen
mit [WS2O2] 2− -Anionen isotype monokline Strukturen ((NH4)2[WS2O2]-Typ, C2/c,
z.B. K2[WS2O2]: a = 1126.5(4), b = 708.2(2), c = 971.5(4) pm, β = 121.86(3) o ) bilden, sind
die K + - und Cs + -Salze dimorph und kristallisieren zusätzlich in den bisher unbekannten
Strukturtypen (P21/c bzw. Pbcn). Die Trithiowolframate von Rb + und Cs + kristallisieren
im orthorhomischen β-K2[SO4]-Typ (Pnma), der auch von den Tetrathiowolframaten gebildet
wird. Am Beispiel der Cs-Dithiooxowolframate wurde der thermisch induzierte Phasenübergang
(bei ca. 100 o C) von der orthorhombischen Struktur (Pbcn, VFormeleinheit =
805 10 6 pm 3 ) in die monokline Form ((NH4)2[WS2O2]-Typ, C2/c, VFormeleinheit = 797
10 6 pm 3 ) thermoanalytisch untersucht. Bei ca. 260 o C tritt unter Volumenzunahme eine
Umwandlung in eine neue Phase im β-K2[SO4]-Typ (Pnma) ein. In den wasserfreien Salzen
sind die [WSxO4−x] 2− -Anionen so gepackt, dass die Oxo-Liganden vier- bis fünffach
durch Kationen koordiniert sind, während die Thio-Liganden Koordinationszahlen von 6
bis 7 aufweisen. Die Anordnung der Wolframattetraederanionen lässt sich auf einfache Metallpackungen
zurückführen. Die Strukturen der Salzhydrate und Ammoniumsalze werden
hinsichtlich ihrer Wasserstoffbrückenstruktur diskutiert. Die Bindungssituation wird anhand
der Zustandsdichten aus FP-LAPW-Bandstrukturrechnungen auf Basis der experimentellen
(W-S)-/(W-O)- Abstände analysiert.
[1] E. Corleis, Liebigs Ann. Chem. 1886, 232, 244-270. [2] G. Krüss, Liebigs Ann. Chem.
1884, 225, 1. [3] K. H. Schmidt, A. Müller Coord. Chem. Rev. 1974, 14, 115. [4] A. Müller,
W. Sievert, Z. Anorg. Allg. Chem. 1974, 403, 251. [5] A. Lehner, K. Kraut, C. Röhr, Acta
Crystallogr. 2010, A66, 186. [6] A. Lehner, C. Röhr, Z. Anorg. Allg. Chem. (in Vorbereitung).
34

Multiple partial melting events in the contact aureole of the Reinfjord ultramafic complex,
Seiland Igneous Province, Northern Norway
Xiaoyan Li, Kurt Bucher
Reinfjord ultramafic complex, one of the largest layered peridotite and gabbro plutons in the
Seiland Igneous Province, emplaced into the lower part of the Sørøy Succession of Kalak nappe
complex, which developed a contact aureole during the prograde metamorphism. The protolith of
material contributing the contact aureole consists of psammite and semipelite, shallow-marine sediment
that deposited on the Baltica continental margin of the Iapetus Ocean. Based on available researches in
the area, we found that multiple thermal events could have changed the lithological unit extensively,
with varying significances. In order to decipher the complicated geological process, we dedicated
abundant field and laboratory works.
Several assemblages have been observed and used as indicators of multiple thermal events:
(1) Sil-Kfs-Ab: layered texture, likely product of breakdown of muscovite.
(2) Bt-Sil-Kfs-Pl-Qtz: different subsets as inclusions in garnets, act as the reactant of biotitebreakdown
incongruent melting reaction, forming peritectic garnet.
(3) Opx-Grt-Sil-Bt: occuring locally, biotite partial melted.
(4) Hc-Crn with or without Sil: as bone-like crystals in the muscovite matrix with the whole patch
enclosed by garnet and biotite, and hercynite encloses sillimanite, likely resulted from back
reaction between minerals and melts.
(5) Ms or Bt: replacing the cordierite and garnet respectively, the new hydrous minerals that formed
by residual melt or fluid reacting with the anhydrous minerals.
35

Redox-Neutral Atom-Economic Rhodium-Catalyzed
Coupling of Terminal Alkynes with Carboxylic Acids
Toward Branched Allylic Esters
Alexandre LUMBROSO, Philipp KOSCHKER, Bernhard BREIT*
Institut fűr Organische Chemie und Biochemie, Freiburg Institute for Advanced Studies (FRIAS),
Albert-Ludwigs-Universität Freiburg, Albertstrasse 21, 79104 Freiburg im Breisgau, Germany
Branched allylic alcohols and their ester derivatives are important building blocks in
organic synthesis. Numerous approaches for their preparation are known, including
asymmetric versions. [1] More recently, formation of allylic esters via CH bond oxidation has
attracted considerable interest and is emerging as a new tool in the synthesis of complex
molecules. [2] A drawback of these reactions with regard to atom economy is the requirement
of stoichiometric amounts of an oxidant. In this work, we have found that using the
diphosphine ligand DPEphos L and [Rh(COD)Cl]2, the addition of carboxylic acids to
terminal alkynes lead to the branched allylic esters B with high chemoselectivity (Scheme
1). [3] The reaction is a redox-neutral formal propargylic CH oxidation with concomitant
hydride shift where the alkyne serves as an internal oxidant to be reduced simultaneously
toward the alkene. This original atom-economic process is applicable to a broad substrate
scope and displays a wide functional group tolerance.
Scheme 1. Addition of carboxylic acids to terminal alkynes via a redox-neutral
atom-economic process
[1] D. M. Hodgson, P. G. Humphreys, In Science of Synthesis: Houben-Weyl Methods of Molecular
Transformations; Clayden, J. P., Ed.; Thieme: Stuttgart, Germany, 2007; Vol. 36, pp 583.
[2] M. S. Chen, N. Prabagaran, N. A. Labenz, M. C. White, J. Am. Chem. Soc. 2005, 127, 6970.
[3] A. Lumbroso, P. Koschker, N. R. Vautravers, B. Breit, J. Am. Chem. Soc. 2011, 133, 2386.
36

Do photosynthetic bacteria have a protective mechanism
against carbonate precipitation at their surfaces?
Raul E. Martinez, Insitute für Geochemie, Universität Freiburg, Albertstr. 23b, D-79104
Freiburg, raul.martinez@minpet.uni-freiburg.de
Closed reactor kinetic experiments, SEM and TEM imaging, EDX analyses, and
zeta potential measurements were used to assess the existence of metabolic process
protecting cyanobacteria against carbonate mineralization on their surfaces. Carbonate
precipitation rates measured at pH of ~8.2 and 23 °C in initially supersaturated solutions
in the presence of active Synechococcus sp. and Planktothrix sp. correspond closely to
those measured in analogous inorganic control experiments. TEM imaging and EDX
analysis indicates the absence of Ca 2+ on active Synechococcus sp. and Planktothrix sp.
surfaces. Electrophoretic measurements of active cyanobacteria surfaces demonstrate
development of a positive surface potential on active Synechococcus sp. and Planktothrix
sp. cyanobacteria at pH 8–10. This positive charge was suppressed by the presence of 1
mM HCO3 - but enhanced by increasing aqueous Ca 2+ concentration in the fluid phase.
These observations suggest the existence of a mechanism, based on the metabolic
maintenance of a positive surface charge at alkaline pH, protecting active cyanobacteria
against Ca 2+ adsorption and subsequent carbonate precipitation on their surfaces. [1]
References:
[1] Martinez, R. E., Gardes, E., Pokrovsky O. S., Schott, J., Oelkers, E. H., Geochim.
Cosmochim. Acta. 2010, 74, 1329.
37

Zn-Indide der Alkalimetalle mit der Anionenstruktur des β-rhomboedrischen
Bors
V. Mihajlov 1 and C. Röhr 1
1 : Institut f. Anorg. u. Analyt. Chemie, Universität Freiburg, Deutschland
viktoria@goethite.chemie.uni-freiburg.de
β-rhomboedrisches Bor, β-B105 (hR, R3m, Z=3), ist eine der allotropen Modifikationen
des elementaren Bors. Die Kristallstruktur besteht aus zwei kristallographisch unterschiedlichen
B12-Ikosaedern im Verhältnis 1:3 und zwei verbrückten kondensierten
Dreifachikosaedern B28, die über exo-Bindungen miteinander verknüpft sind (Abb. 1).
Sie stellt eine hierarchische Variante des MgCu2-Typs dar, bei der die Ikosaeder die
Cu- und die Dreifachikosaeder die Mg-Plätze einnehmen ([B28]2B12[B12)3]). Bereits die
ersten Einkristallstrukturdaten [1] zeigen, dass entsprechend der Wade-Regeln nicht
alle Borlagen der B28-Einheit voll besetzt sind.
Analoge Gallide und Indide existieren aus sterischen Gründen nur mit Alkalimetallkationen
A + in den Lücken zwischen den Clustern, z.B. Na26K8Ga99.1 [2] oder K14Na20In96.3
[3]. Die Elektronenbilanz wird entsprechend durch stärker ausgedehnte Defekte im Anionenteilgitter
wiederhergestellt [4].
Ein Ausgleich des durch die zusätzlichen A-Atomen eingebrachten Elektronenüberschusses
kann durch den Einbau von elektronenärmeren Elementen der späten Übergangsmetallreihe
erreicht werden, z.B. in K34Zn18.8In86.2 [5]. Ausgehend von dieser
defektfreien Phase wurde der sterische Einfluss der Kationengröße auf die Struktur
systematisch durch Austausch der Alkalimetalle röntgenographisch anhand von Einkristalldaten
untersucht. Das Zn/In-Verhältnis bleibt dabei annähernd gleich, die Valenzelektronenkonzentration
(vec) pro M beträgt 3.16 und liegt damit im engen Bereich
zwischen 3.10 und 3.38, in den alle Verbindungen mit diesem Strukturtyp auftreten.
Berechnungen der elektronischen Strukturen mittels FP-LAPW-DFT-Methoden zeigen
in Übereinstimmung hiermit ein deutlich ausgeprägtes Minimum in der totalen
Zustandsdichte am Ferminiveau.
Die sechs Kationenpositionen können in zwei Gruppen unterteilt werden. Die drei Positionen
A(1,2,3) (insgesamt 14 Atome) weisen eine kleinere Koordinationssphäre mit
der Koordinationszahl (CN) 12 (gekappte Tetraeder) und die Positionen A(4,5,6) (insgesamt
20 Atome) eine größere mit einer CN von 15 bzw. 16 auf.
Ausgehend von K34Zn18.8In86.3 führte der sukzessive Austausch von K + durch das kleinere
Kation Na + zu der Grenzzusammensetzung K14Na20Zn16.7In88.3 (a = 1771.44(5),
c = 3890.7(2) pm), bei der alle A(1,2,3)-Lagen mit Na + und A(4,5,6)-Lagen mit K +
besetzt sind.
Umgekehrt erfolgte der Austausch von K + auf den Lagen A(4,5,6) durch größere Kationen
ebenfalls nahezu vollständig, so dass Rb13.3K20.7Zn18.9In86.1 und Cs12.3K21.7Zn19.2In85.8
(a = 1838.30(5)/1846.27(9), c = 4017.97(2)/4039.7(2) pm) die Randverbindungen darstellen.
Als Bildungsvoraussetzung für den vorgestellten Strukturtyp können somit zwei limitierende
Größen genommen werden: Zum einen der bereits erwähnte enge vec-Bereich
und zum anderen ein Kationen/Anionen-Radienverhältnis von rA/rM = 0.97 - 1.14 für
die Lagen A(1) bis A(3) bzw. rA/rM = 1.14 - 1.31 für A(4) bis A(6).
[1] J. L. Hoard, D. B. Sullenger, C. H. L. Kennard, R. E. Hughes J. Solid State Chem. 1, 268-277 (1970).
[2] C. Belin, M. Charbonnel, J. Solid State Chem. 64, 57-66 (1986).
[3] B. Li, J. D. Corbett, J. Am. Chem. Soc. 127, 926-932 (2005).
[4] C. Belin, M. Charbonnel, Coord. Chem. Rev. 178-180, 529-564 (1998).
[5] B. Li, J. D. Corbett, Inorg. Chem. 45, 8958-8964 (2006).
38

C
B
A
c
O
a
b
Abbildung 1:
39
2
Zn
In

Liposomal encapsulation of arsenic trioxide for effective treatment of
neuroblastoma cancer
Müller, I. 1 , Schleicher, S.², Schubert, R. 1
1
Lehrstuhl für Pharmazeutische Technologie und Biopharmazie,
Universität Freiburg
²Klinik für Kinder und Jugendmedizin (Hämotologie und Onkologie),
Universitätsklinikum Tübingen
Arsenic trioxide (ATO) is effectively used as an antineoplastic drug 1 for the treatment
of various cancer diseases, for example acute promyelotic leukemia and certain
neuroblastoma forms. Due to side effects like renal and heart failure that might occur
in oral application, a liposomal encapsulation of ATO for parenteral administration is
recommended.
This allows for increased accumulation of ATO in the tumor environment because of
the EPR effect (enhanced permeability and retention) in tumor-supporting blood
vessels carrying gap junctions. The accumulation effect could be stimulated by an
active targeting of these liposomes towards specific antigens being expressed on
cancer cells’ surfaces. This may be achieved via the combination of a sterol based
anchor integrated in the liposomal bilayer and corresponding antibodies covalently
attached to this anchor. It was already shown that a sole incorporation of ATO
solution in liposomes did not result in a stable formulation (Kallintieri et al 2004) 2 .
Therefore a remote loading mechanism using nickel acetate (Chen et al. 2006) 3 was
adapted that enabled a precipitation and thus entrapment of ATO inside of the
hydrophilic liposomal core.
Here preliminary results with regard to cell viability of certain neuroblastoma cell
lines are shown. Cell viability results upon treatment with ATO-solution and
liposomal entrapped ATO, respectively, are compared to solely nickel-loaded or
unloaded control liposomes, as well as non-treated control cells.
1 Ferrara et al. : Acute promyelocytic leukemia: what are the treatment options?
Expert Opin Pharmacother. 2010 Mar;11(4):587-96
2 Kallintieri et al.: Arsenic trioxide liposomes: encapsulation efficiency and in vitro stability, J Liposome Res. 2004; 14(1-2):27-
38.
3 Chen et al. : Lipid encapsulation of arsenic trioxide attenuates cytotoxicity and allows for controlled anticancer
drug release, J. Am. Chem. Soc., 2006, 128 (41), 13348-13349
40

CONVERTING AN AROMATIC TO A POLYKETIDE:
APPLICATION TO THE SYNTHESIS OF THE
C 1 -C 11 FRAGMENT OF RIMOCIDIN
Luc Nachbauer and Reinhard Brückner*
Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität,
Albertstraße 21, D-79104 Freiburg; lucnachbauer@hotmail.com
Rimocidin (1) is a member of the polyol/polyene family of macrolide antibiotics, which consists
of ~200 naturally occurring molecules. We have developed a novel synthesis of the synthetic
equivalent 2 of the northern moiety of rimocidin (1). [1] Our route is based on a straightforward
generation of the polyketide chain by the oxidative cleavage of an ortho-bridged dihydroaromatic
system. [2]
HO
HOOC
14
HO
O
11
18
OH
O-D-mycosamin
1
O OH
nPr
1
O
O
S
S
11
OTBS
S S
Specifically, we synthesized the non-racemic indane 4 from commercially available 5-hydroxyindane
(3) in 5 steps. A tandem reaction consisting of a Birch reduction and ozonolysis led to triketo-ester
5. S,S-acetal formations and stereoselective reductions gave the desired C 1 -C 11 fragment 2.
References:
[1] Isolation: J. W. Davisson, F. W. Tanner, Jr., A. C. Finlay, I. A. Solomons, Antibiot. Chemother. 1951, 1,
289-290.� Total syntheses: G. K. Packard, Y. Hu, A. Vescovi, S. D. Rychnovsky, Angew. Chem. 2004, 116,
2882-2886; Angew. Chem., Int. Ed. 2004, 43, 2822-2826; A. B. Smith III, M. A. Foley, S. Dong, A. Orbin, J.
Org. Chem. 2009, 74, 5987-6001.
[2] Ozonolysis of bridged hydroaromatics: A. J. Birch, F. Fitton, D. C. C. Smith, D. E. Steere, S. R. Stelfox, J.
Chem. Soc. 1963, 2209-2216.� C. L. Kirkemo, J. D. White, J. Org. Chem. 1985, 50, 1316-1319.
41
2
Et
OTBS
1
OTBS

Noxa/Mcl-1 balance decides the effect of the proteasome
inhibitor MG-132 in combination with anticancer agents in
pancreatic cancer cell lines
Katrin Naumann a , Kathrin Schmich a , Christoph Jaeger a , Felix Kratz b , Irmgard Merfort a
a Department of Pharmaceutical Biology and Biotechnology, University of Freiburg, Stefan-
Meier-Str. 19, D-79104 Freiburg, Germany
b Tumor Biology Center, Clinical Research, Breisacher Str. 117, D-79106 Freiburg, Germany
Abstract
Pancreatic cancer progresses aggressively and due to chemoresistance responds poorly to
chemotherapy. Thus, there is an urgent need to understand the mechanisms of cancer cell
resistance in order to generate effective strategies to circumvent intrinsic chemoresistance in
this tumor indication. In this work, three pancreatic cancer cell lines, MIA PaCa-2, MDA
Panc-3, and AsPC-1, were treated with the proteasome inhibitor MG-132 together with
camptothecin, doxorubicin or paclitaxel. The combination of MG-132 and camptothecin in a
ratio of 5:1 gave the most promising results and enhanced cytotoxicity compared to the single
compounds in MIA PaCa-2 cells. The increase is shown to be due to an enhanced caspase-3
activity resulting in apoptosis. Moreover, this combination upregulated the levels of the pro-
apoptotic protein Noxa and reduced the levels of the anti-apoptotic protein Mcl-1. In contrast,
the combination of MG-132 with doxorubicin only resulted in an increased cytotoxic, but also
in a decreased apoptotic effect. The lack of the enhanced apoptosis induction could be
correlated with high levels of Mcl-1 in response to the combined treatment with MG-132 and
doxorubicin.
Thus, the results indicate that regulation of the anti- and pro-apoptotic Bcl-2 family members
Noxa and Mcl-1 predicts the effectiveness of the combination of MG-132 with different
anticancer agents on apoptosis induction in pancreatic cancer cells.
42

Incorporation of the H + -ATPsynthase from E. coli into a planar lipid bilayer and
observation by spFRET
P. Oswald * and P.Gräber *
* Albert-Ludwigs-University of Freiburg, Department of Physical Chemistry, Albertstr. 23a,
79104 Freiburg, Germany
peter.oswald@physchem.uni-freiburg.de
H + -ATPsynthases catalyze the synthesis of ATP from ADP and inorganic phosphate by a
rotational mechanism. Transmembrane proton translocation leads to a rotational movement of
the intramembrane c10-subunits and this movement induces conformational changes at the
catalytic sites which finally lead to ATP synthesis. This rotation has been observed by single
pair Fluorescence Resonance Energy Transfer (spFRET) using double labeled enzymes, which
were integrated into the membrane of liposomes [1]. Catalysis was initiated by an acid-based
transition which generates a transmembrane electrochemical potential difference of protons. In
this work we try to incorporate a double labeled H + -ATPsynthase into a planar lipid bilayer
and we ask whether it is possible to initiate rotation by applying an electric potential
difference across this bilayer. First the double labeled enzyme was reconstituted into
liposomes which contain nystatin and ergosterol. Fusion of the liposome with the bilayer can
be observed by a current which was due the formation of K + -ion channels by nystatin and
ergosterol in the bilayer.
[1] Diez M., Zimmermann B., Börsch M., Schweinberger E., Steigmiller S., Reuter R.,
Felekyan S., Kudryavtsev V., Seidel C., Gräber P., Proton-powered subunit rotation in single
membrane-bound F0F1-ATP synthase, Nat. Struct. Mol. Biol., 11, 135-141, (2004).
43

Chasing electrons – Time-resolved spectroscopy on blue-light
sensitive flavoproteins
Bernd Paulus, Erik Schleicher and Stefan Weber
Albert‐Ludwigs‐Universität Freiburg, Institut für Physikalische Chemie
Albertstraße 21, 79104 Freiburg / Germany
Proteins of the photolyase/cryptochrome‐family use blue light to perform important tasks, such as
entrainment of circadian rhythms, DNA repair, regulation of plant growth and possibly
magnetoreception in migratory birds [1‐3]. To harvest the light and use it for protein function, they
carry a flavin cofactor, e.g. flavin adenine dinucleotide (FAD). The cofactor has to be in the proper
redox state. This is obtained via photoactivation, a process involving the transient formation of
radical pairs [4]. Using time‐resolved optical and EPR‐spectroscopy we study the dynamics of this
process as well as the influence of the surrounding protein environment in different proteins of the
photolyase/cryptochrome‐family.
The work presented has been performed in collaboration with
K. Hitomi and E.D. Getzoff (Scribbs Research Institute, La Jolla, CA), P.J. Hore, A. Robinson, K. Henbest,
K. Maeda and C. Timmel (Oxford University), and A.R. Marion and J.R. Norris (University of Chigaco).
[1] T. Biskup, E. Schleicher, A. Okafuji, G. Link, K. Hitomi, E.D. Getzoff, S. Weber, Angew. Chem. Int.
Ed. 48 (2009) 404‐407.
[2] S. Weber, T. Biskup, A. Okafuji, A.R. Marino, T. Berthold, G. Link, K. Hitomi, E.D. Getzoff, E.
Schleicher, J.R. Norris, J. Phys. Chem. B 114 (2010) 14745‐14754.
[3] K. Henbest, K. Maeda, P.J. Hore, M. Joshi, A. Bacher, R. Bittl, S. Weber, C. Timmel, E. Schleicher,
Proc. Natl. Acad. Sci. USA 105 (2008) 14395–14399.
[4] S. Weber, Biochim. Biophys. Acta 1707 (2005) 1–23.
44

The Deslongchamps Reaction as an Access to the
Dihydroagarofuran Sesquiterpenoids
Denis Petrović and Reinhard Brückner*
Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität, Albertstr. 21,
79104 Freiburg, Germany; E-mail: denispetrovic@gmx.de
The 1300 plants of the Celastraceae family 1 are indigenous to the tropical and subtropical
regions of the world. Their biological effects are noteworthy and can be traced back to a
number of unusually highly oxygenated sesquiterpenoids. 2 Two of them are the
dihydroagarofurans 1 and 2.
Considering 1 and 2 as synthetic targets we plan to derive them from the cis-configured
hexalindione 6 via the densely functionalized octalindione 3. Hexalindione 6 should be
accessible by a DESLONGCHAMPS annulation 3 of the acceptor-substituted enolate 5 � which
results from an in-situ deprotonation of a so-called NAZAROV reagent 4 � to the estersubstituted
cyclohexenone 4. The latter is an activated quinonemonoketal 4. Engaging this
particular functionality in DESLONGCHAMPS annulations represents one of several extensions
of the scope of this versatile reaction. 5 In the case at hand, choosing 4 furnishes allows to
obtain a hexalindione (6) rather than an octalindione, which would result otherwise. The
latter, of course, would lack one of the C=C bonds and thereby a site poised for further
elaboration.
1 R. Brüning and H. Wagner, Phytochemistry 1978, 17, 1821-1858.
2 A. C. Spivey, M. Weston, S. Woodhead, Chem. Soc. Rev. 2002, 31, 43-59.
3 P.-Y. Caron.; P. Deslongchamps Org. Lett.2010, 12, 508-511 and literature cited therein.
4 I. N. Nazarov and S. I. Zauyalou. Zh. Obshch. Khim. 1953, 23, 1703-1706.
5 J. F. Lavalée, P. Deslongchamps, Tetrahedron Lett. 1988, 29, 5117-5118. For our earliest contribution see T.
Tricotet, R. Brückner, Eur. J. Org. Chem. 2007, 1069-1074.
45

Inter- and Intramolecular Hydroacylation of Alkenes
employing a Novel Bifunctionnal Catalyst System
Damien Regent, Nicolas Vautravers, Bernhard Breit*
Institut für Organische Chemie und Biochemie, Freiburg Institute for Advanced Study (FRIAS),
Albert-Ludwigs-Universität Freiburg, Albertstr. 21, 79104 Freiburg, Germany
e-mail: bernhard.breit@chemie.uni-freiburg.de
The hydroacylation of alkenes is an attractive atom-economic [1] synthetic method for the preparation of
ketones from aldehydes via transition metal-catalyzed C-H bond activation. [2] One drawback of this
reaction is the decarbonylation of the intermediate acyl-metal hydride complex (Scheme 1), which
lead to a side product. Many methods have thus been developed in order to suppress it. [3] Inspired by
the work of Jun et al [4] , we optimized a new bifunctional ligand L combining an aminopicoline moiety
and a phosphine function (Scheme 1). [5]
R 1
O
+ H R2
metal binding
substrate binding
The aminopicoline moiety would act as a reversibly
bound directing group allowing for facile C-H activation
while simultaneously preventing the undesired
decarbonylation through formation of an aldimine.
Integration of the phosphine function would not only
form an active hydroacylation catalyst, but also enhance
the binding constant of the directing group to the
catalytically active rhodium center.
We applied successfully our bifunctionnal ligand to the inter- and intramolecular hydroacylation of
alkenes [5] , with yields up to 83% in the intermolecular hydroacylation of substituted benzaldehydes 1
and diverses alkenes 2 (Scheme 2).
Intermolecular Hydroacylation of Alkenes
O
[Rh(COD) 2]BF4 (5 mol%)
FGR1 H
+ R FG
2
ligand L (5 mol%)
toluene (c = 1.1 M)
150 °C, 1 h
1
FG R1
2 3
Scheme 2
9 exemples
up to 93% yield
Similarly, excellent yields up to 98% were also reached in the intramolecular reaction of substituted ovinylbenzaldehyde
4 yielding valuable 1-indanone derivatives 5 (Scheme 3).
References:
Ph 2P
ligand L
N N
Rh
H R2 R 1
#
R 1
Intramolecular Hydroacylation of Alkenes
[Rh(COD) 2]BF4 (5 mol%)
FGR H
ligand L (5 mol%)
O
toluene (c = 1.1 M)
150 °C, 1 h
O
Scheme 1
R 2
4 5
Scheme 3
10 exemples
up to 98% yield
[1] B. M. Trost, Science 1991, 254, 1471-1477; B. M. Trost, Acc. Chem. Res. 2002, 35, 695-705.
[2] M. C. Willis, Chem. Rev. 2010, 110, 725-748.
[3] T. B. Marder, D. C. Roe, D. Milstein, Organometallics 1988, 7, 1451-1453; T. Kondo, N. Hiraishi, Y.
Morisaki, K. Wada, Y. Watanabe, T. Mitsudo, Organometallics 1998, 17, 2131-2134; A. A. Roy, C. P.
Lenges, M. Brookhart, J. Am. Chem. Soc. 2007, 129, 2082-2093.
[4] Y. J. Park, J.-W. Park, C.-H. Jun, Acc. Chem. Res. 2008, 41, 222-234; C.-H. Jun, E.-A. Jo, Eur. J.
Org. Chem. 2006, 2504-2507
[5] N. R. Vautravers, D. D. Regent, B. Breit, Chem. Comm. 2011, ASAP
46
FG R
O
O
R 2 FG

SYNTHESIS AND TESTING OF 2,7-NAPHTHYRIDONES AS A NOVEL
CLASS OF SIRTUIN INHIBITORS
Schiedel, M. 1 , Schemies, J. 1 , Jung, M. 1
1 Institute of Pharmaceutical Sciences, Albert-Ludwigs-Universität Freiburg,
Germany, http://jungm.de
Sirtuins represent a specific NAD + -dependent class of histone deacetylases
(HDACs). By using NAD + as a cofactor, these enzymes cleave off the acetyl
groups from the ε-amino group of lysines in histones and other proteins, e.g. p53,
FOXO proteins, p300 or HIV tat. The human family of sirtuins consists of seven
members, which are distributed to different cell compartments and involved in the
regulation of various physiological processes like apoptosis, cell differentiation,
metabolism, DNA recombination and HIV tat transactivation. Thus, sirtuin
inhibitors are interesting potential drugs for drug discovery. [1]
Besides the deacetylated lysines, the products of the catalytic reaction are 2`-Oacetyl-adenosine
diphosphate ribose and nicotinamide, which itself acts as a
physiological sirtuin inhibitor. Thus, we chose nicotinamide as a lead structure for
synthesis of new potential sirtuin inhibitors.
Synthesis of 2,7-naphthyridones [2]
Quaternary N 1 -alkylnicotinamides were treated with an alkaline solution of
methylketones. After a nucleophilic addition of the deprotonated methylketone
compound to the pyridinium salts in the 4-position, the 2,7-naphthyridones were
formed by dehydrogenation. As an oxidant formic acid was used. The highly
fluorescent products were either isolated as a dipolar ion out of the aqueous layer
or its hydrochloride out of an ethanolic phase.
N +
R
O
NH 2
+
C
H 3
O
R'
KOH
0°C
O
R'
N
R
O
NH 2
N
HCOOH
100°C OH - H +
In-vitro testing
To discover new sirtuin inhibitors among the synthesized 2,7-naphthyridones, we
have used a screening assay, which is based on fluorescence polarisation. [3] As the
most potent derivative, we could identify 2-benzyl-8-hydroxy-6-(naphthalene-2yl)-2,7-naphthyridin-2-iumchloride
(structure shown below) with 52% inhibition of
Sirt2 at a concentration of 25 µM.
N +
Cl
References:
[1] Trapp J., Jung M. (2006) Curr Drug Targets, 7, 1553-1560
[2] Palfreyman M.N., Wooldridge K.R.H. (1974) J. Chem. Soc., Perkin Trans, 1,
57-61
[3] Milne J.C., Lambert P.D. et al. (2007) Nature, 450, 712-716
47
OH
N
N +
R
R'
+ H +
N +
R
R'
N
O

Quantenchemische und Matrixisolationsuntersuchungen
zu Eisentetrafluorid
T. Schlöder, S. Riedel*
Institut für Anorganische und Analytische Chemie, Albert-Ludwigs Universität Freiburg, Albertstr. 21, D-79104
Freiburg i. Br. (Germany)
tobias.schloeder@cpg.uni-freiburg.de, sriedel@psichem.de
Eisendi- und trifluorid sind stabile Verbindungen, welche sowohl experimentell 1, 2 als auch
theoretisch 3, 4 gut untersucht sind. Das nächst höhere Eisenfluorid FeF4 hingegen ist schwerer
zu greifen. Eine neue IR Bande in einem Matrixisolationsexperiment wurde diesem Molekül
zugeordnet, nachdem FeF3 mit dem Fluoratomdonor CeF4 zur Reaktion gebracht wurde;
jedoch konnten im Massenspektrum kein entsprechendes Signal gefunden werden 5 .
Hier möchten wir über quantenchemische Rechnungen zu Strukturen und
Schwingungsspektren der Eisenfluoride auf bis zu CCSD(T) Niveau berichten. Weiterhin
wurde eine thermochemische Stabilitätsanalyse unternommen, bei der die Elimination von
Fluoratomen und -molekülen (uni- und bimolekular) in Betracht gezogen wurde. Nachdem
das Ergebnis zeigte, dass FeF4 in der Gasphase stabil ist, wurden Matrixislationsexperimente
zur Synthese durchgeführt. Dabei wurde eine Mischung aus CeF4 und FeF3 erhitzt und die
gasförmigen Spezies über der Probe zusammen mit den Wirtsgasen Neon oder Argon
eingefroren. Die IR Spektren zeigen zusätzlich zu den Banden von FeF2 und FeF3 eine weitere
Absorption, welche basierend auf den quantenchemischen Rechnungen dem Eisentetrafluorid
zugeordnet werden kann.
1. Vogt, N., Journal of Molecular Structure 2001, 570, (1-3), 189-95.
2. Hargittai, M., The Journal of Chemical Physics 2005, 123, (19), 196101-2.
3. Solomonik, V. G.; Stanton, J. F.; Boggs, J. E., The Journal of Chemical Physics 2005,
122, (9), 094322-12.
4. Solomonik, V. G.; Stanton, J. F.; Boggs, J. E., The Journal of Chemical Physics 2008,
128, (24), 244104-9.
5. Rau, J. V.; Nunziante, C. S.; Chilingarov, N. S.; Leskiv, M. S.; Balducci, G.; Sidorov,
L. N., Inorg. Chem. Commun. 2003, 6, (6), 643-5.
48

Hochkonzentrierte, fließfähige Kolloiddispersionen
Schneider, René, rene.schneider@physchem.uni-freiburg.de
Wiemann, Malte, malte.wiemann@uni-freiburg.de
Bartsch, Eckhard, eckhard.bartsch@physchem.uni-freiburg.de
Hochkonzentrierte, fließfähige kolloidale Dispersionen besitzen ein breites Anwendungs­
spektrum in Industrie und Alltag. Es konnte an einem Modellsystem harter Kugeln,
bestehend aus Polystyrol(PS)­Mikrogelkolloiden in einem organischen Lösemittel,
mittels Lichtstreuung gezeigt werden, dass durch Zugabe von freiem, nicht
adsorbierendem Polymer der Bereich der Fließfähigkeit zu höherem Volumenanteil der
Kolloide verschoben werden kann [1,2]. Die Wahl des Modellsystems war durch die
Anforderung der Lichtstreuung bestimmt, dass die hochkonzentrierten Dispersionen
transparent sein müssen (Isorefraktivität, d.h. Brechungsindex Kolloid ≅ Brechungs­
index Medium). Um zu erkennen, ob die erhaltenen Ergebnisse allgemein gelten, wurde
ein neues System synthetisiert, welches dem Modell harter Kugeln näher kommt. Des
weiteren stellte sich die Frage, ob dieser Effekt auch für die in der Anwendung wichtige
Materialklasse der wässrigen Polymerdispersionen auftritt und genutzt werden kann.
Daher war die Entwicklung eines weiteren Modellsystems erforderlich, das
Isorefraktivität in Wasser gewährleistet. Dazu wurden selbst synthetisierte Per­
fluoroacrylat­Partikel charakterisiert, die über eine Schicht aus Polyethylenoxidketten in
Wasser sterisch stabilisiert werden.
Links: Phasendiagramm
einer PS-Mikrogel/PS-
Polymer-Mischung [2]. C P
gibt die Polymerkonzentration
an, ϕ den
Kolloidvolumenbruch. fluid
= fließfähige Dispersionen.
Rechts: Hochkonzentrierte
Fluoracrylat-Dispersion in
Wasser (n D =1.332; rechts)
ϕ
und in Wasser/Glyzerin-
Mischung (n 1.37; links)
D
Referenzen:
[1] Eckert, T. und Bartsch, Phys. Rev. Lett. 89, 1257 (2002)
[2] Eckert, T. und Bartsch, J. Phys.: Condens. Matter 16, S4937 (2004)
49

Albert-Ludwigs-Universität Freiburg
Specific siRNA delivery to Alveolar Rhabdomyosarcoma by
co-modified liposomes
Doris Sehi, Regine Süss
Department of Pharmaceutical Technology and Biopharmacy, University of Freiburg,
Germany
Tel: +49 761 203 6329, email: doris.sehi@pharmazie.uni-freiburg.de
Rhabdomyosarcoma is the most frequent malignant soft tissue tumor in
childhood. Especially the more aggressive subtype named Alveolar RMS
(ARMS) is highly resistant to all forms of therapeutical treatment that are
currently available [1]. RNA interference offers a new promising approach for
the treatment of ARMS but lacks an effective way to deliver the siRNA into
the cancer cells. This study aims to develop a liposomal formulation which is
able to protect the siRNA from degradation on its way to the tumor and to
achieve specific interaction and successful uptake when having arrived there.
Therefore, surface-modified SPC/Cholesterol liposomes were designed
carrying the RGD peptide to mediate selective interaction with
rhabdomyosarcoma cells and the TAT peptide to promote effective uptake. In
vitro experiments with the co-modified liposomes show promising results.
1. Parham, D., Ellison, D., (2006) Rhabdomyosarcomas in Adults and Children, An
Update. Arch Phathol Lab Med 130, 1454-1465
2. Ko, YT. et al (2009) Gene delivery into ischemic myocardium by double-targeted
lipoplexes with anti-myosin antibody and TAT-peptide Gene Therapy 16, 52-59
50

CoRS Curator: A Text Curation and Annotation Tool
Senger C*, Erxleben A, Grüning BA, Günther S
Pharmaceutical Bioinformatics,
Institute of Pharmaceutical Sciences, University of Freiburg, Germany
*e-mail: christian.senger[at]pharmazie.uni-freiburg.de
Introduction: Compound related information is usually published in text-form, which is not very
well structured. Furthermore, associated information is usually scattered over a great number of
articles. The Compound Research System (CoRS) is a DFG supported project for establishing an
automated molecule research system and associated modules to extract and connect relevant
information. CoRS Curator is a software tool for facilitating selection, care, and presentation of
objects in texts and for transferring this information to databases in a standardised form.
Methods: CoRS Curator uses publicly available text-mining and information retrieval software to
preselect texts and highlights found entities. Users are enabled to annotate pieces of texts with
predefined entities via text-highlighting, allowing for a high degree of traceability to the
information's origin.
In a test case, researchers used the tool to collect information on a set of more than 6000 abstracts
related to the compound class of sesquiterpene lactones, enabling an iterative further development
and evaluation of the software in terms of interrater agreement (agreement of curators on
annotations of text pieces), curation speed, and annotation outcome.
Results: The curators needed ~90 man-hours to curate 6650 abstracts for 530 compounds. About
78% of the abstracts contained redundant information. Interrater agreement was 72% on a test set.
More than 2800 pieces of text were annotated with entities (e.g. “source organism“, “assay“, or
“therapeutic action“) providing important information of the analysed compounds.
51

Tag der Forschung - 05.07.2011
Improving transfection efficiencies of lipid-mediated gene delivery
A. Steinbach, R. Süss
- Influence of the lipid/DNA complex preparation technique -
Institute of Pharmaceutical Sciences, Department of Pharmaceutical Technology and Biopharmacy,
Sonnenstr. 5, D-79104 Freiburg;
annette.steinbach@pharmazie.uni-freiburg.de
Lipid/DNA complexes are commonly used non-viral gene delivery systems bearing great
potential due to their low host-immunogenicity. To date their clinical use is restricted by
limited transfection efficiencies caused by cellular barriers [1].
To explain and improve low transfection efficiencies of DC 30/DNA complexes, different
stages of the transfection process were addressed as potential transfection barriers in the
two previously characterized cellular models A-10 SMC and MDCK II [2]. Particular
attention was paid to complex internalisation, intracellular processing, nuclear uptake of
complex-released pDNA as well as transgene expression.
DC 30/DNA complexes derived from two preparation protocols involving treatment of
starter liposomes with a sonication tip before complexation with pDNA compared to an
untreated preparation were included in our studies.
Transfection deficiency of DC 30/DNA complexes in MDCK II cells could not be related to
one specific step in the cellular transfection process. The entirety and interplay of all
cellular processes from internalisation to transgene expression need to be considered to
contribute to the low transfection efficiency in some cellular systems. The preparation
protocol of DC 30/DNA complexes was found to have an influence on cellular uptake and
cell line dependently on the transfection efficiency, but no impact on intracellular
processing or nuclear uptake of complex released pDNA could be observed.
References:
1. Hama et al., Quantitative and mechanism-based investigation of post-nuclear delivery events between
adenovirus and lipoplex. Nucleic Acids Research, 2007. 35(5): p. 1533-1543
2. Schneider S., Dissertation University of Freiburg, 2010
52

MEGABLOCKS IN THE RIES IMPACT CRATER, GERMANY: NEW DISCOVERIES AND
STATISTICAL ANALYSIS OF DISTRIBUTION AND LITHOLOGIES.
S. Sturm1, W. Willmes1, H. Hiesinger1, T. Kenkmann2 and G. Pösges³, 1Institut für Planetologie, Westfälische
Wilhelms-Universität Münster, Germany (s.sturm@uni-muenster.de), 2 Institut für Geowissenschaften,
Universität Freiburg, Germany, 3Rieskrater-Museum Nördlingen, Germany.
The Ries crater is a well-preserved complex crater, 25 km in diameter, located in Bavaria, southern Germany [1
and references therein]. The impact occurred in a heterogeneous target that consisted of ~600 m sedimentary
cover (Triassic to Jurassic) resting over a crystalline basement [2]. It consists of (i) a 10-12 km diameter inner
crater basin filled with suevite and post-impact lake deposits, (ii) a collar zone of upturned and overturned highly
faulted and shock metamorphosed material called the “inner ring” and (iii) a zone of large blocks (“megablock
zone”) that were displaced during the crater formation process [e.g., 3]. The Ries has been intensively studied,
however the megablock zone still poses questions regarding crater formation mechanics. Here we present new
data of the megablock zone using a combined approach of remote sensing analysis and shallow drillings.
Formation of megablocks is caused by two processes: (i) They were either thrown outwards during the crater
excavation stage and were deposited simultaneously with the continuous ejecta blanket (Bunte breccia) and/or
(ii) slumped inwards during the modification stage, leading to a complex juxtaposition of allochthonous
crystalline and allochthonous and parautochthonous sedimentary megablocks. Their abundance and distribution
is a tribute to the pre-impact geology, the impact process and post-impact erosion. The megablock zone exhibits
a hummocky morphology that is dominated by large Malmian and crystalline megablocks easily detectable at the
surface [2, 4]. Mega-blocks are also present in the subsurface as they have been buried by Suevite, Bunte Breccia
or post-impact sediment deposition [5]. These subsurface megablocks have only rarely been detected so far and
are not shown in the present geological map of the Ries [6].
References: [1] Stöffler, D. (1977) Geologica Bavaria, 75, 443-458. [2] Pohl et al. (1977) Impact and Explosion Cratering,
Flagstaff. [3] Wünnemann, K. et al. (2005) In: Large Meteorite Impacts III, Geological Society of America, Boulder. [4]
Hüttner and Schmidt-Kaler (1999) Geologica Bavarica, 104, 7-76. [5] Collins et al. (2008), Meteoritics and Planetary
Science, 43, 12, 1955-1977. [6] Geologische Karte des Rieses 1:50000 (2005) Bayerisches Geologisches Landesamt,
München.
53

Overproduction of Aquifex aeolicus complex I in E. coli.
M. Vranas, and T. Friedrich
Institute for Organic Chemistry and Biochemistry,
Albert-Ludwigs-University Freiburg, Germany
The NADH:ubiquinone oxidoreductase, also called complex I, is the main entrance
point of electrons into the respiratory chains. Within the complex electrons are
transferred from NADH to Ubiquinone (Q) coupled with a proton translocation across
the membrane according to:
NADH + Q + 5H + → NAD + + QH2 + 4H +
The enzyme comprises a noncovalently bound flavin mononucleotide and several ironsulfur
clusters as co-factors. In Aquifex aeolicus, a hyperthermophilic bacterium with an
optimal growth temperature of 95ºC, the 13 nuo-genes, coding for NuoA-N subunits of
complex I, are organized in three different loci [1, 2].
Heterologous production of A. aeolicus complex I is attempted in different Escherichia
coli strains. In some strains the chromosomal nuo-operon was deleted. The deletion of
the 16kb nuo-operon was performed by Lambda-Red Recombineering technique, a
PCR mediated gene replacement method [3, 4].
Expression plasmids were constructed containing various groups of the A. aeolicus
nuo-genes, including the entire nuo-operon from the three loci mentioned above. The
strains were transformed with the expression plasmids and the heterologous
overproduced proteins were isolated and characterized.
1. Kohlstadt, M. et al., Heterologous Production, Isolation, Characterization and Crystallization
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54

Structural Deformations in the Central Pit of a Martin
Crater as an Indicator for Impact Direction
G. Wulf, M. H. Poelchau and T. Kenkmann,
Institute of Geosciences – Geology, Albert-Ludwigs-University Freiburg, Germany (gerwin.wulf@geologie.uni-freiburg.de)
The majority of impacts on planetary bodies occur at an oblique impact angle to the target
surface because the incidence angle follows a Gaussian probability distribution with a mean
value of 45° [1]. The ejecta blanket is the most distinctive indicator for the impact direction in
oblique impacts, showing the formation of “forbidden” zones and “butterfly” patterns [2,3].
The position of the central uplift relative to the crater center was proposed as another diagnostic
feature [4], but its statistical relevance could not be verified thus far [5] as the position
might be caused by heterogeneous target structures [6]. Another promising indicator for
obliquity is provided by the internal structure of central uplifts. Some terrestrial craters show a
preferred stacking of layered bedrock in the central uplift [7,8] with bedding striking perpendicularily
to the long axis of the crater ellipse [8]. The interpretation that this imbrication is
caused by remnant horizontal momentum transferred from the impacting projectile to the target
during an oblique impact [8] is supported by three-dimensional numerical simulations [9].
Further analysis of Martian craters [10,11] provide evidence that preferred strike orientation
in the central uplift could be indicative for an impact direction. Here we present new results of
the mapping of an unnamed Martian crater extending and confirming these results for a central
pit structure of an oblique impact crater.
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[2] Gault D. E. and Wedekind J. A. (1978) Proc. Lunar Planet. Sci. Conf. 9: 3843-75.
[3] Anderson, J. L. B. et al. (2004) Meteoritics & Planet. Sci., 39, 303–320.
[4] Schultz, P. H. and Anderson, R. R. (1996) Geol. Soc. of Amer. Spec. Paper, 302, 397-417.
[5] Ekholm, G. A. and Melosh, H. J. (2001) Geophy. Res. Let., 28, 623-626.
[6] Collins, G. S. et al. (2008) Earth & Planet. Sci. Let., 270, 221–230.
[7] Scherler, D. et al. (2006) Earth & Planet. Sci. Let. 248, 43-53.
[8] Kenkmann, T. and Poelchau, M. H. (2009) Geology, 37, 459-462.
[9] Shuvalov, V. V. et al. (2004) Meteoritics & Planet. Sci., 39, 467-479.
[10] Wulf, G. et al. (2011) LPSC, 42, #1440.
[11] Poelchau, M. H. et al. (2009) LPSC, 40, #1796.
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