CAREER: NMR Insights into the Influence of Dynamics on SH3 Domain Mediated Protein Interactions

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The objective of this project is to develop novel techniques to characterize Nuclear Magnetic Resonance (NMR) relaxation of proteins in the solution state. The development and implementation of these techniques will be facilitated by (i) advances in theoretical methods for experimental design and analysis, (ii) major improvements in commercially available instrumentation, and (iii) the development of cost-effective methods for the bacterial expression and purification of labeled proteins. These methods will be applied to study protein dynamics over a wide range of timescales, with special emphasis on the characterization of protein sidechain dynamics since sidechains play a central role in defining protein-ligand interactions and hence physiological function. These methods will alleviate many of the deficiencies of current experiments by (i) providing information on the details of local dynamics on the fast, ps-ns timescale, (ii) allowing complete characterization of slow, correlated motions on the microsecond-ms timescale, and (iii) being applicable to larger proteins and protein complexes. The PI will utilize molecular dynamics simulations in conjunction with theoretical modeling to identify specific dynamic modes that contribute to the measured relaxation rates. This combination of experimental and computational techniques will allow the identification of key functional modes and the changes thereof on protein interactions. The PI will apply this methodology to elucidate the dynamic determinants of the intra- and inter-molecular interactions involving two cytosolic components of the NADPH oxidase complex - p47phox and p67phox. A very important structural module, the SH3 domain plays a central role in defining protein interactions in these cases. Thus, information gleaned from these studies will provide a self-consistent, space-time view of SH3 domain-mediated protein interactions.The PI will place special emphasis on recruiting students from various backgrounds and at various levels of sophistication, undergraduate and graduate, into his research program. He will redesign undergraduate courses to provide students with the knowledge of cutting edge biophysical techniques and develop a new course on NMR spectroscopy targeted towards advanced undergraduates and graduate students. This course will introduce students to both the theoretical as well as the applied aspects of modern biomolecular NMR spectroscopy both through lectures as well as a series of "hands-on" workshops. The PI currently participates, and will continue to do so, in science outreach programs such as ACS-SEED. This program is designed to facilitate underprivileged high-school students to gain research experience over the summer. CUNY City College has a large minority population. Thus the research and teaching components of this project will encourage those sections of society underrepresented in the sciences to pursue scientific careers both in academia and in industry. Facilitating minority participation in the sciences through teaching, mentoring, career counseling and participating in outreach programs for disadvantaged students is a key goal of this CAREER project.