High-resolution tracking of microtubule motility driven by a single kinesin motor.
Kinesin is a microtubule-based motor protein that contains two identical force-generating subunits. The kinesin binding sites along the microtubule lie 8 nm apart (the dimension of the tubulin dimer), which implies that kinesin must translocate a minimum distance of 8 nm per hydrolysis cycle. Measurements of kinesin's microtubule-stimulated ATPase activity (approximately 20 ATP per sec) and velocity of transport (approximately 0.6 micron/sec), however, suggest that the net distance moved per ATP (approximately 30 nm) may be greater than one tubulin dimer under zero load conditions. To explore how kinesin translocates during its ATPase cycle, we constructed a microscope capable of tracking movement with 1-nm resolution at a bandwidth of 200 Hz and used this device to examine microtubule movement driven by a single kinesin motor. Regular stepwise movements were not observed in displacement traces of moving microtubules, although Brownian forces acting on elastic elements within the kinesin motor precluded detection of steps that were < 12 nm. Though individual steps of approximately 16 nm were occasionally observed, their infrequent occurrence suggests that kinesin rarely moves abruptly by distances of two or more tubulin subunits during its ATP hydrolysis cycle. Instead it is more likely that kinesin moves forward by the distance of only a single tubulin subunit under zero load conditions.
Malik
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Neuroscience Graduate Program, University of California, San Francisco 94143.
Brillinger
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Vale
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D000255:Adenosine Triphosphate / Q000031:analogs & derivatives; D000465:Algorithms; D000818:Animals; D001921:Brain; D002417:Cattle; D049832:Decapodiformes; D005583:Fourier Analysis; D016547:Kinesins / Q000737:chemistry* / Q000502:physiology*; D007700:Kinetics; D008853:Microscopy / Q000295:instrumentation / Q000379:methods; D008863:Microspheres; D008870:Microtubules / Q000737:chemistry* / Q000502:physiology*; D009899:Optic Lobe, Nonmammalian; D013816:Thermodynamics; D013997:Time Factors
C029273:P(3)-1-(2-nitro)phenylethyladenosine 5'-triphosphate; D000255:Adenosine Triphosphate; D016547:Kinesins