T4 LYSOZYME AS A MODEL FOR PROTEIN STABILITY

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DESCRIPTION: This proposal is directed towards obtaining a better understanding of the relationships between protein structure, stability and dynamics using T4 lysozyme as a model system. The proposal presents four specific aims: 1) The folding pathway connecting the folded and unfolded states will be investigated, with specific emphasis on the structure and dynamics of an early kinetic intermediate and equilibrium states that appear related to it. Approaches are described to monitor the relative rates of encounter of various parts of the polypeptide chain using an excimer probe.; 2) The mechanism of coupling between different parts of the sequence, that fold as an apparent unit in the wild-type protein but become uncoupled in certain mutants. Pulsed hydrogen exchange methods will be employed to define the rates at which different parts of the protein attain native-like structure.; 3) The accessibility of water and small hydrophobic ligands to cavities in the core of T4 lysozyme will be examined.; 4) A major effort will be undertaken to develop the methods necessary to observe and interpret the effects of the application of mechanical forces designed to unfold small assemblies of proteins attached to the tip of an atomic force microscope. These types of measurements will provide a new perspective of the energy landscape of proteins.