Role of three dimensional structure of proteins in signal transduction

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The aim of the present research was to elucidate the roles of proteins in signal transduction process on the structural basis. We also developed the protein expression and purification systems for preparation of amount of proteins required for structural studies. In the course of the present research project, a number of protein structures were determined by either NMR spectroscopy or X-ray crystallography. Grb2 n-SH3 and SH2 domains complexed with proline rich peptide from Sos and phosphotyrosine containing peptide from Shc, respectively were determined and the general mechanism for recognition of target sequences were elucidated. Three dimensional structure of destrin was determined by NMR. Although the sequence homology between destrin family and gelsolin family proteins was not present, the three dimensional structure of both proteins were very similar, suggesting the same mechanism for actin severing. Thus, the general mechanism for severing of the actin fiber was proposed. The three dimensional structure of midkine was also determined by NMR and elucidated the mechanism of dimer formation on heparin oligosaccharides. The dimer formation of midkine seems to be essential for activation of the receptor and its biological function. We also determined the CRD domain of PKC and suggested the mechanism of DAG recognition. By X-ray crystallography, a number of protein structures were determined including transmitter and receiver domains of bacterial signal transduction system. We also made protein expression and purification system for N-WASP, Tob and CAF1. N-WASP is essential for formation of philopodia in response to the growth factor receptor signal. Tob and CAF1 play pivotal role in cell cycling. The structural studies started for both proteins.