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 role of the cotranslationally active chaperone Hsp70 in modulating polypeptide conformation and folding is studied. Investigations are primarily carried out by multidimensional nuclear magnetic resonance to reveal high-resolution information. In living cells, kinetic arguments indicate that there is ample time for conformational sampling to take place cotranslationally, before synthesis of the full-length polypeptide chain has been completed. The vectorial nature of protein synthesis and the complex cellular environment potentially affect de novo protein folding. It is therefore important to characterize how proteins fold in the cellular context. Molecular chaperones are known to interact with nascent ribosome-bound polypeptide chains during protein synthesis. However, it is not known to what extent they affect polypeptide conformation, and whether or not they are able to reshape the cotranslational and immediately post-translational folding landscapes. The current work specifically addresses this issue. The effect of complex formation on polypeptide conformation is investigated in an in vitro model system comprising the substrate binding domain of the cotranslationally active Hsp70 chaperone and its peptide substrates. The changes in the substrate binding domain upon peptide binding, as well as the modulation of substrate conformation upon chaperone-binding are addressed. The study provides insights at the residue level on the conformational changes associated with complex formation involving the Hsp70 chaperone.
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