Long-term Objectives: 1. To explore new types of motility within the myosin superfamily; 2. To address fundamental unresolved issues in motor protein biochemistry such as the relation of the powerstroke to the actin binding transitions and product release steps of myosin. Significance: Motility and contractility are essential for almost all biological processes. Detailed quantitative knowledge of the enzymology of molecular motors is a prerequisite for understanding biological motility and the basis of physiological processes connected to, among others, cardiac function and malfunction, asthma, axonal regeneration and sensory illnesses.
Specific Aims : 1. We will test the hypothesis that mechanical load has a functionally important effect on the product release kinetics of non-muscle myosin II; 2. We will assess if myosin X, a membrane-associated motor, has a unique mechanism of action that differs from all described types of myosin-based motility; 3. By mutational perturbation of the product release steps of myosin V, we will investigate the relation of product release to the mechanical step (powerstroke) and actin binding transitions of myosin. We will also establish the correspondence between the kinetic effects of the mutations and motility and processivity. Research Design and Methods: We will address the above questions by using single- and double-headed recombinant myosin constructs for solution kinetic, spectroscopic, biochemical and molecular mechanical investigations.
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