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Mark T. Fisher
Associate Professor |
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Biochemists are now aware that protein folding inside the cell is, in some cases, assisted by other essential proteins called molecular chaperones. Work in our laboratory focuses on elucidating the molecular mechanism of action for one family of molecular chaperones called the chaperonins. The large oligomer chaperonin-60 (Cpn-60), binds some nascent protein folding intermediates and facilitates their folding and assembly reactions. Chaperonin function requires ATP and is modulated through the interaction of a smaller second accessory oligomeric chaperonin protein called chaperonin-10 (Cpn-10). Using Escherichia coli dodecameric glutamine synthetase (GS) as our model substrate, we have determined; 1) that chaperonins facilitate the formation of assembly competent GS monomers and 2) the driving force for chaperonin-assisted folding and assembly of GS comes from the binding free energies of ATP and cpn-10. Current work in our laboratory focuses on determining the thermodynamic parameters (such as binding free energies, enthalpies, and entropies) for the various chaperonin-protein interactions. Numerous investigators have demonstrated that the solution requirements are variable for a number of protein substrates. Our hypothesis simply states that the different requirements are dictated by differences in initial binding free energies of protein substrates for chaperonin-60. By comparing the energetics and mechanisms of chaperonin interactions with a number of different protein substrates, we will obtain a more comprehensive explanation for chaperonin action. The techniques that we apply in our work include rapid scanning uv-visible second derivation spectroscopy, microcalorimetric titration, stopped flow fluorescence and absorption spectroscopies, and general protein chemistry. Our long term goal is to define the specific properties that govern chaperonin protein-protein substrate interactions in the cell. Determining the role that chaperonins play in insuring proper protein folding at the molecular level will lead to the design of highly efficient protein production systems that will mass produce biomedically relevant protein products.
