The goal of this research is to understand the mechanisms by which a protein folds spontaneously to a unique 3-dimensional conformation. Presumably, all the information required for folding is contained in the primary amino acid sequence. Thus, changes at key positions in the sequence could have profound effects on folding, which can be identified and used to elucidate the nature of the folding process. Dihydrofolate reductase from E.coli is the model protein chosen for study. Its gene has been cloned and its 3-dimensional structure is known. Mutants will be generated by mutagenesis directed at sites believed to be important in the folding reactions. Equilibrium and kinetic studies will be carried out on the folding/unfolding transition in wild type and mutant proteins using ultraviolet, fluorescence, circular dichroism and NMR spectroscopy. The results will be fit to simple models, and conclusions drawn regarding the structural features that are critical to folding. From such studies the rules that govern folding may eventually emerge, so that it will become possible to make predictions on the function of a protein based on its primary structure.
