Objective
The calcium signal is involved in many cell events. It is deciphered through the binding to calcium binding proteins, which undergo conformational changes and interact with specific target proteins or supramolecular complexes. The goal of the project is to compare the calcium binding mechanisms with and without target structures for three members of the EF-hand super-family: calmodulin, calcium-vector protein, and sarcoplasmic calcium-binding protein of Nereis, in order to understand the molecular mechanism of theirs specific cellular actions.
The cooperative folding units and flexibility in these proteins and their complexes with target proteins
(i), the calcium binding mechanism for them in a medium mimicking the cytoplasmic environment ;
(ii), and the modulation of the calcium binding mechanism upon interaction with target structures;
(iii) will be characterized.
These data will allow understanding how such an interaction modulates the activity of the target structures. The interaction between calmodulin and microtubule associated proteins (MAPs) will be particularly considered with its consequences on microtubular architecture. Finally, the effects of some inhibitors with a potentiality in cancer on the interactions of MAPs with tubulin and calmodulin according to calcium binding steps will be revealed. The used techniques include differential scanning and isothermal titration calorimetry, mass-spectrometry, NMR, multifrequency phase fluorometry, fluorescence correlation spectroscopy, flow dialysis, and site-specific mutagenesis.
The unique combination of techniques and available calcium-binding proteins must allow to precisely correlate the cationic sites occupancy with the flexibility and dynamic changes of the folding units of calcium binding proteins and, therefore, with theirs interaction with target structures. In addition, knowledge will be gained on the structural and thermodynamic characterization of these complexes and such studies will be the starting point for the design and screening of new classes of more specific calmodulin inhibitors. In return, these inhibitors will be valuable tools to settle the controversy over the role of calmodulin in the regulation of the microtubular system. Such knowledge will allow deciphering the role of calcium binding proteins in the cellular filaments regulation (microtubules, myosin filaments). The project is an extension of the previous INTAS grants 94-2068 which resulted in six joint publications including two in Biochemistry (1997,1998).
Topic(s)
Call for proposal
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13385 Marseille
France