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Biologische Systeme und Medizin Poster: Mi., 14:00–16:30 M-P194
Lysozyme and Staphylococcal Nuclease solutions studied by small-angle xray
scattering
Christof Krywka 1 , Christian Sternemann 1 , Nadeem Javid 2 , Roland
Winter 2 , Metin Tolan 1
1 Fachbereich Physik, DELTA, Universität Dortmund, D-44221 Dortmund –
2 Fachbereich Physikalische Chemie, Universität Dortmund, D-44221 Dortmund
Ever since x-rays were employed for structure determination, small-angle x-ray scattering
(SAXS) has retained its unique position among methods that allow to investigate
structure on an Angstrom length scale. It is left with no competition when structure
of partially or completely disordered particles is sought and its major advantage is the
feasibility of in situ measurements, e.g., on protein molecules dissolved in their natural,
aqueous environment at low concentration.
The biological activity, thermodynamic stability along with the physical and chemical
properties of proteins are strongly influenced by the structure of their aqueous microenvironment.
Hence, understanding the effects of various types of cosolvents typical for
cellular conditions on the structure and dynamics of proteins is crucial for a deeper
insight into protein stability, folding, aggregation and fibrillation processes. The latter
play an important role in many conformational diseases, such as Alzheimer, Huntington,
Creutzfeldt-Jakob, and Parkinson. It was shown that the latter processes are in
fact markedly influenced by the type and concentration of cosolvents.
As a part of the human immune system, lysozyme breaks carbohydrate chains, destroying
the structural integrity of bacteria cell wall and leading them to burst under their
high internal osmotic pressure. The influence of the cosolvents tetrafluoroethylene,
sodium chloride, and glycerol on the native state structure of lysozyme was studied using
SAXS measurements on aqueous solutions of lysozyme at beamline BL9 of DELTA
synchrotron, University of Dortmund. A wide range of concentrations of pure lysozyme
as well as lysozyme in the presence of cosolvents, typical for conditions where unfolding
sets in, was probed. For the higher concentrated samples, information about the intermolecular
interaction potential could be obtained from analysis of the intermolecular
structure factor.
The biological function of staphylococcal nuclease is to catalyze the hydrolysis of the
phosphate backbone of DNA and RNA. Unlike lysozyme, this protein can fold and
unfold reversibly due to the lack of disulfide bonds or free sulfhydryl groups. This
peculiarity opens up the possibility to studying the intermediate states of unfolding
next to its native and denaturated state by applying a reversible condition that induces
unfolding or refolding, such as temperature or pressure. For this reason, SAXS
measurements were also performed on staphylococcal nuclease in a wide pressure and
temperature range in the absence and presence of various cosolvents, using a home built
high pressure cell, and subsequently analysis of the tertiary structure of the different
conformational states was performed.