STRUCTURE-FUNCTION STUDIES OF 70 KD HEAT SHOCK PROTEINS

70千道尔顿热休克相关蛋白（hsp70相关蛋白）是广泛的

The 70 kilodalton heat shock-related proteins (HSP70-related) are wide- spread and essential for cell viability. They appear to be intimately involved the processes of protein folding, targeting and maturation. Bacterial and parasite HSP70-related proteins are highly immunogenic; host immune responses to these highly conserved proteins may lead to autoimmunity syndromes. This proposal focuses on the question: What are the biochemical functions of the HSP70-related proteins in cells? The general strategy is structure- function studies at the molecular level. The specific aims are: 1.Elucidate the ATPase mechanism using x-ray crystallography and site- specific mutagenesis. 2.Search for evidence for or against a major conformational change in the proteins using small-angle solution x-ray scattering. 3.Delimit the substrate binding functions within the substrate recognition domain of the bovine heat shock cognate protein (HSC70), using clathrin as a substrate, by constructing (i) C-terminal deletion mutants and (ii) proteins which are chimeras of HSC70 and another HSP70 related protein, BiP. 4.Probe the functional significance of the structural and sequence similarities of the ATPase fragment and actin, using site-specific mutagenesis in conjunction with ATPase and clathrin uncoating activity assays. 5.Examine the feasibility of using Laue diffraction on the HSC70 ATPase fragment to determine the structures of the reaction intermediated during catalysis. 6.Attempt to crystallize intact HSP70-related proteins and/or their substrate recognition domains.