Kinetics: a tool to study molecular motors.
Molecular motors are enzymes that couple the energy from nucleoside triphosphate hydrolysis to movement along a filament lattice. The three cytoskeletal motor superfamilies include myosin, dynein, and kinesin. However, in the last decade it has become apparent that the nucleic acid-based enzymes (DNA and RNA polymerases as well as the DNA helicases) share a number of mechanistic features in common with the microtubule and actin motors despite the fact that their cellular functions are so different. This review addresses the mechanistic approaches that have been used to study molecular motors. We discuss the basic biochemical techniques used to characterize a protein preparation, including active site determination and steady-state kinetics. In addition, we present the transient-state kinetic approaches used to define a mechanochemical cycle. We attempt to integrate the information obtained from kinetic studies within the context of motility results to provide a better understanding of the contribution of each approach for dissecting unidirectional force generation.
Gilbert
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SP
Department of Biological Sciences, University of Pittsburgh, 518 Langley Hall, Pittsburgh, Pennsylvania 15260, USA. spg1@pitt.edu
Mackey
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D000255:Adenosine Triphosphate / Q000378:metabolism; D001665:Binding Sites; D001703:Biophysics / Q000379:methods*; D002458:Cell Fractionation; D003402:Creatine Kinase / Q000378:metabolism; D019281:Dimerization; D029721:Drosophila Proteins; D017022:Flow Injection Analysis; D016547:Kinesins / Q000378:metabolism*; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008870:Microtubules / Q000378:metabolism*; D008956:Models, Chemical; D020409:Molecular Motor Proteins / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D010710:Phosphates / Q000032:analysis; D010725:Phosphocreatine / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary
D029721:Drosophila Proteins; D020409:Molecular Motor Proteins; D010710:Phosphates; C066696:ncd protein, Drosophila; D010725:Phosphocreatine; D009243:NAD; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase; D003402:Creatine Kinase; D016547:Kinesins