MECHANICS OF KINESIN--MICROTUBULE-BASED MOTOR PROTEIN

点击翻译按钮获取中文摘要

The long-term objective of the proposed studies is to understand the physical chemistry of chemomechanical transduction, the conversion of chemical energy contained in high-energy phosphate bonds to mechanical energy used to power intracellular movement. This process is accomplished by several enzymes, termed motor proteins, that include myosin from muscle cells, dynein from cilia and flagella, and cytoplasmic dynein and kinesin from eukaryotic cells in general. Current models, such as the crossbridge model, postulate that the underlying transduction process is a cyclic reaction of a motor molecule with a cytoskeletal polymer, an actin filament in the case of myosin and a microtubule in the instances of dynein and kinesin. After binding to the filament, the motor protein is thought to undergo a conformational change, the power stroke, that produces an increment of movement. The protein then releases the filament before rebinding at another site along the filament and initiating another cycle. The specific aim of the proposed experiments is to test directly such models by making mechanical measurements of the transduction reaction at the single-molecular level. The movement of microtubules across surfaces coated at low density with purified kinesin will be visualized by dark- field microscopy. Special apparatus, previously used by the investigator to study force-sensitive ion channels in hair cells of the ear, will be used to exert forces and to measure displacements with subnanometer precision on a millisecond timescale. The distance that a single kinesin molecule moves a microtubule upon the hydrolysis of a molecule of ATP will be determined. After characterizing the movement of microtubules by single kinesin molecules, the nature of the interactions between several kinesin molecules moving one microtubules by single kinesin molecules, the nature of the interactions between several kinesin molecules moving one microtubule will be studied in order to predict the behavior of large assemblies of motor proteins such as those found in muscles and cilia. Because of the structural and biochemical similarities between kinesin and other motor proteins, the elucidation of the molecular events underlying transduction by kinesin should significantly increase the understanding of cellular motility in general. It is hoped that this understanding may lead to more rational treatments of muscle disorders such as heart disease, or to better methods of selectively interfering with pathological cellular movements such as the invasion and proliferation of tumor cells, and the transport of viruses between the cell membrane and the nucleus.