WIDE ANGLE SCATTERING OF PROTEIN SOLUTIONS FOR FOLD DETERMINATION

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Wide-angle x-ray scattering from proteins in solution (WAXS) provides a set of capabilities for analysis of the structure of proteins not accessible by other methods. Our experiments at BioCAT are designed to develop those capabilities. We are currently pursuing three avenues of research: (i) Assignment of fold on the basis of WAXS patterns: We are compiling a database of WAXS patterns from proteins of known and unknown folds with the goal of using it as a basis for fold assignment for proteins that cannot be crystallized. WAXS data from over 100 proteins of known structure have been collected thusfar. (ii) Protein-ligand interactions: WAXS provides the capability of studying structural changes in proteins induced by binding of small molecule ligands. Ligand-induced structural changes are too large to be accommodated within a crystal lattice, making the development of alternative approaches critical. We have collected data on 10 proteins in the presence and absence of small molecule ligands, and carried out detailed analysis of the structural changes induced for 5 of these systems. (iii) Protein breathing: In comparing WAXS patterns from dilute protein solutions with those from concentrated solutions, we discovered systematic differences that could be attributed to structural fluctuations that are much greater in dilute solution. Detailed analysis of these patterns is being used to obtain a quantitative measure of the scale of the fluctuations. Suppression of these fluctuations in concentrated solution appears to be due to molecular crowding, a phenomena that promotes protein stability in the cell cytoplasm.