Structure and Function of Molecular Motors

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DESCRIPTION (provided by applicant): We will extend our studies of the mechanism by which chemical energy is transformed into mechanical energy by motors from the myosin and kinesin families. The approach for the kinesin motors will build on our previous work which determined the structures of several of the microtubule based motors in multiple conformations, and which used spectroscopic methods to determine how these structures changed during the cycle. We will obtain structures for new members of the kinesin family. We will investigate the conformational changes that occur in these structures by obtaining dynamic information using spectroscopic probes, both fluorescent and paramagnetic. We will expand our knowledge of the neckdocking mechanism of conventional kinesin. We will initiate studies of NCD, which may work through a mechanism very different from kinesin. Myosin VI is a processive motor with a long step, but a short lever arm, and spectroscopic probes will be used to investigate this unusual motor. In addition we will investigate the structure and function of both the motor region and the cargo-binding region of a member of another motor family, dynein. These structural and functional data will be complemented by measurements of motor mechanics. Oligomers of the actin filament and their complexes with myosin fragments will be formed, and their properties and structures determined. We will initiate a new direction for our program, and investigate the cargo domains of the motor proteins and identify their protein partners that together connect the motors to their loads. The cargo domains are less well characterized than are the motor domains, and only a few of their many partners are known. We will employ chemical, structural, biochemical, and genetic techniques to identify the proteins involved in cargo recognition, determine structures for selected cargo binding domains and their partners, and characterize their function. In summary, our goal of understanding the mechanism of biological motors requires: a) determination of their atomic resolution structures, b) knowledge of how these structures change in specific states, and c) measurements of the mechanics and energetics associated with these states. In this program we have brought together a unique group of investigators that can extend our knowledge in each of these areas leading to a better picture of the molecular mechanism of these two motors that produce force and motion.