AnnualReport2020

Life & Health Sciences 2020
50
Dr Vishwanatha
Thimmalapura Marulappa
LE STUDIUM / Marie Skłodowska-Curie
Research Fellow
Smart Loire Valley General Programme
From: University Medical College Groningen - NL
In residence at: Center for Molecular
Biophysics (CBM) - Orléans
Nationality: Indian
Dates: February 2019 to May 2020
I have joined the Prof. Sureshbabu’s laboratory
(Bangalore University, India) in July 2009 for a PhD
programme. The main objectives of my research
work were the design and synthesis of a novel class
of peptidomimetics. After finished my PhD in 2014,
Alexander Dömling (University of Groningen, The
Netherlands) offered a postdoc position to work on
multicomponent reactions. I am very fortunate to have
had an additional experience to work on radiochemistry
laboratory at the medical college under the guidance
of Prof. Elsinga. I have committed for the opportunity
to expand my ideas and past research activities for
the synthesis of complex structures such as peptides
and proteins. I worked as a postdoc in Dr Aucagne’s
group on drug discovery and organosulfur-based
peptidomimetic synthesis. I received several award
grants such as CSIR senior research fellowship, travel
grants to attend the 2018 International Symposium on
Chemical Biology in Switzerland and 2019 American
Peptide symposium in the USA. With the help of
Studium fellowship, I could success in getting an exiting
project at Ledien University Medical Center.
Dr Vincent Aucagne
Host Scientist
Vincent Aucagne received his PhD from the
University of Orléans (2002), working with
Patrick Rollin on the development of synthetic
methodologies to elaborate carbohydrate mimics.
Following post-doctoral research with Prof. David
Leigh at the University of Edinburgh (2003-2006) in
the field of mechanically-interlocked architectures
and molecular machines, he returned to Orléans
to join the CNRS Center for Molecular Biophysics
(CBM), as a CNRS Chargé de Recherche (2006) in
the group of Dr Agnès Delmas. He currently holds a
Director de Recherche position, leads the “Synthetic
Proteins and Biorthogonal Chemistry” research
group, and is the coordinator of the “Molecular,
Structural and Chemical Biology” team. His current
research interests focus on the development of
synthetic methodologies for the chemical synthesis
of proteins for application to the deciphering
of biological processes at the molecular level.
DEVELOPMENT OF NOVEL CHEMOSELECTIVE
LIGATION TECHNIQUES FOR PROTEIN SYNTHESIS
The production of proteins by chemical synthesis is a very promising
alternative to biotechnological techniques for applications to the
deciphering of biological mechanisms at the molecular level, drug
discovery and synthetic biology. It is particularly useful for accessing
site-specifically modified proteins. Current technologies focus on the
modular assembly of unprotected peptide fragments through highly
chemoselective reactions called “chemical ligations”.
This approach revolutionised the field about thirty years ago, but there
is still only a very few reactions compatible with this purpose available
to date. The overall goal of the project is to develop novel ligation
reactions for chemical protein synthesis. In particular, the chemical
ligation of peptide thioacids with N-activated peptides (imidazolyl
ureas) has been investigated, with the goal to transform a known
non-chemoselective reaction (carboxylic acid/imidazoylurea coupling
develloped by Campagne) into a chemo and regio-selective reaction
compatible with aqueous environments typically used for the ligation
of unprotected peptides.
The rational behind the idea to replace carboxylic acids by thioacids is
that, under acidic conditions, thioacids are reactive whereas side chain
functional groups in the peptides such as amines and carboxylic acids
are expected to be unreactives.
The ligation reaction between peptide thioacids and imidazolyl urea
peptides involves three key challenges:
1. synthesis of unprotected peptide thioacids
2. synthesis of imidazolyl urea peptides and
3. ligation under aqueous conditions.
First, I investigated the synthesis of unprotected peptide thioacids
which are notably difficult to prepare, and rather unstable. A model
peptide was synthesized having a C-terminal hydroxy-benzyl cysteine
group at the C-terminus (so called crypto-thioester) as described
previously in the host laboratory. After the elongation on solid support,
the peptide was released in solution and purified through HPLC. This
crypto-thioester peptide was treated with trimethoxy benzyl thiol
(Tmob-SH) leading to the formation of trimethoxy benzyl thioester
peptide which was purified by HPLC. Very conveniently, trimethoxy
benzyl thioester peptide is efficiently converted into peptide thioacid by
a simple acidic treatment prior to use in ligation reactions. The second
challenge was the synthesis of imidazolyl urea peptides. For this,
activation of amine was carried out on solid support by the treatment
with carbonyl diimidazole.
The imidazolyl urea activated peptide was then released from resin. I
systematically studied the stability of model imidazolyl urea peptide
in aqueous conditions. It was found that, the imidazolyl urea activated
peptides are reasonably stable in water at acidic pH, thus compatible
with the reaction with peptide thioacids.
Next we have carried out typical ligation reaction between unprotected
peptide thioacid derived from LYYRG and unprotected Imidazolyl urea
peptide was taken as simple alanine methyl ester under acidic pH
based on a previously synthesized dipeptide Boc-Phe-Ala-OMe as a
solid proof of concept.
LE STUDIUM CONFERENCES
CHALLENGES AND PROSPECTS IN CHEMOSELECTIVE
LIGATIONS: FROM PROTEIN SYNTHESIS TO SITE-
SPECIFIC CONJUGATION
The main objectives are the
interface between chemistry and
biology, especially methods to
synthesize proteins by chemical
way and recombinant fashion.
Additionally, by using organic
chemistry reactions, how proteins
will be modified and used them
as biological tools to study the
protein function.
The conference was really exiting
and many world-renowned
professors presented their
ongoing discoveries and future
prospects. Many oral and poster
presentations have been selected
from the Ph.D. and postdocs
across Europe. We acknowledge
Studium and many pharma companies including European peptide symposia
for their generous financial assistance. During the conference, many new
techniques have been learnt for the protein chemical synthesis and new
chemistries for the bioconjugation of proteins and their applications to solve
the biological complexity.
Many world-renowned scientists are attended and presented their discoveries
which are relevant to chemistry and biology. Key scientists are Prof. Lutz
Ackermann, Dr Didier Boturyn, Dr Fabienne Burlina, Prof. Alexander Dömling,
Prof. Beat Fierz, Prof. Matthew B. Francis, Dr Sébastien Gouin, Prof. Wenshe
Liu, Dr Oleg Melnyk, Dr Cyrille Sabot, Dr Olivier Sénèque, Dr Denis Servent, Dr
Frédéric Taran, Prof. Alesia Tietze, Dr Vladimir Torbeev, Prof. Carlo Unverzagt,
Dr Birgit Wiltschi,
Life & Health Sciences 2020
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