ayout 1 - EMBL Grenoble

EMBL Research at a Glance 2009
Klaus Scheffzek
PhD 1995, Max-Planck-
Institut für Medizinische
Forschung, Heidelberg.
Postdoctoral research at the
Max-Planck-Institut für
Molekulare Physiologie,
Dortmund.
Team leader at EMBL since
1999.
Signal transduction – disease proteins
Previous and current research
in a segment which represents only 10% of the protein and remains the
only clearly defined biochemical function of the protein.
We are following a structural proteomics approach to explore possible
functions of the remaining 90% of the protein. The idea is to identify
neurofibromin segments that can be expressed as soluble proteins, determine
the structures of such fragments, and by comparison with
known protein structures or by bound ligands obtain ideas for functional/biochemical
experiments. Work on this project offers the opportunity
to contribute to a challenging and physiologically exciting
research topic. Our main technique is X-ray crystallography, with
other methods of protein characterisation being increasingly employed.
Using this approach, we have recently discovered a novel bipartite
module containing a lipid binding Sec14-homology (NF1-Sec)
and a previously undetected pleckstrin homology (NF1-PH)-like domain,
binding cellular glycerophospholipids.
Future projects and goals
Defects in signalling pathways are often associated with the occurrence of severe diseases, with
cancer being a very common example. We are interested in understanding the mechanisms of
pathogenesis associated with cancer-related diseases. In previous work we have characterised the
regulation of Ras, a GTP binding protein mutated in 30% of human tumours, and the related Rho
proteins. Ras functions like a binary molecular switch cycling between GTP-bound ‘ON’- and
GDP-bound ‘OFF’-states; Ras mediated GTP hydrolysis turns the switch off. This intrinsically
slow process is enhanced by so-called GTPase activating proteins (GAPs). Oncogenic Ras mutants
are permanently activated and are not sensitive to GAPs. In earlier studies we have elucidated
the chemical mechanism of GTPase activation and explained why oncogenic Ras mutants
are not GAP sensitive.
Currently a major focus is on neurofibromatosis type 1 (NF1), a genetic disease with an incidence
of 1 in 3,500 newborns. NF1 patients have an increased tumour risk, may show a variety of developmental
defects and frequently have learning disabilities. The NF1 gene encodes a huge protein
(20 times larger than the oxygen carrier protein myoglobin), termed neurofibromin, and when
mutated is responsible for the pathogenesis of the disease. Neurofibromin acts as a Ras specific
GAP, and in some tumour types lacking the protein, Ras is indeed hyperactive. The GAP activity
of neurofibromin resides
Structure of a bipartite module from neurofibromin composed of
a Sec14 homologous (NF1-Sec) and a pleckstrin homology
(NF1-PH) like domain bound to a cellular glycerophospholipid.
A major goal is to arrive at a 3D model of neurofibromin. In addition to the ‘divide and conquer’ strategy, we are gradually returning to the
‘conquer only’ approach by trying to overexpress the full-length neurofibromin in various eukaryotic hosts. With its availability we will also
consider electron microscopy to study its structure. We are increasingly including automated strategies to identify soluble protein fragments
that are accessible to biochemical/structural analysis. In addition, we will continue searching for interaction partners of neurofibromin and
investigate their role for the function of the protein. Studying Sec14- like domains in the context of other signal regulatory proteins such as
RhoGAPs, RhoGEFs and PTPases will be an important direction in the future.
Further projects of the laboratory include signalling by eukaryotic and prokaryotic protein kinases, novel phosphoryltransfer systems, structural
neurobiology and the regulation of retroviral transcription.
Selected references
Hothorn, M., Neumann, H., Lenherr, E.D., Wehner, M., Rybin, V.,
Hassa, P.O., Uttenweiler, A., Reinhardt, M., Schmidt, A., Seiler, J.,
Ladurner, A.G., Herrmann, C., Scheffzek, K., Mayer, A. (2009).
Catalytic core of a membrane-associated eukaryotic polyphosphate
polymerase. Science, in press
Anand, K., Schulte, A., Vogel-Bachmayr, K., Scheffzek, K. & Geyer,
M. (2008). Structural insights into the cyclin T1-Tat-TAR RNA
transcription activation complex from EIAV. Nat. Struct. Mol. Biol.,
15, 1287-1292
Welti, S., Fraterman, S., D’Angelo, I., Wilm, M. & Scheffzek, K.
(2007). The sec1 homology module of neurofibromin binds cellular
glycerophospholipids: mass spectrometry and structure of a lipid
complex. J. Mol. Biol., 366, 551-562
D’Angelo, I., Welti, S., Bonneau, F. & Scheffzek, K. (2006). A novel
bipartite phospholipid-binding module in the neurofibromatosis type
1 protein. EMBO Rep., 7, 17-179
50