This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.The protein NSF and its relative p97 are large hexameric ATPases responsible for disassembling protein complexes. NSF disassembles the coiled-coil complex of SNARE proteins that mediate bilayer fusion during intracellular vesicle transport and exocytosis. The related enzyme p97 removes misfolded proteins from the endoplasmic reticulum membrane. The mechanism by which the energy of ATP hydrolysis is transduced into the work performed on substrates is not understood. We have previously used SAXS to obtain solution structures of p97 in four nucleotide states: ATP, ADP+AlFx, ADP, and no nucleotide, at a nominal resolution of 8.6 . These structures reveal significant conformational changes as this protein goes through the hydrolytic cycle. We propose to 1) extend these studies to NSF, and 2) perform time-resolved studies on both p97 and NSF in order to obtain direct kinetic data on the conformational changes that can be correlated with the chemical hydrolysis of ATP.
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