Substeps within the 8-nm step of the ATPase cycle of single kinesin molecules

Abstract

Kinesin is a molecular motor that moves processively by regular 8-nm steps along microtubules. The processivity of this movement is explained by a hand-over-hand model in which the two heads of kinesin work in a coordinated manner. One head remains bound to the microtubule while the other steps from the alphabeta-tubulin dimer behind the attached head to the dimer in front. The overall movement is 8 nm per ATPase cycle. To investigate elementary processes within the 8-nm step, we have developed a new assay that resolves nanometre displacements of single kinesin molecules with microsecond accuracy. Our data show that the 8-nm step can be resolved into fast and slow substeps, each corresponding to a displacement of approximately 4 nm. The substeps are most probably generated by structural changes in one head of kinesin, leading to rectified forward thermal motions of the partner head. It is also possible that the kinesin steps along the 4-nm repeat of tubulin monomers.

Kinesin: a molecular motor with a spring in its step

“…Sub-8 nm steps similar to those in curve B in figure 5a have been observed experimentally by Coppin et al (1996) using a 110 Hz low-pass filter. However, more recent measurements on kinesin by Nishiyama et al (2001) seem to show that the second sub-step may occur within ca. 50 µs of the first.…”

“…Figure 5a shows a Monte Carlo simulation (Thomas & Thornhill 1998) Nishiyama et al (2001). Curve A is the unfiltered signal sampled at 100 kHz, whilst curves B and C show filtered data as in (a).…”

“…We show that the integration of this step into the kinesin cycle (Hackney 1994;Duke & Leibler 1996;Hancock & Howard 1999;Astumian & Derenyi 1999;Schnitzer et al 2000) produces a scheme that can account quantitatively for the kinesin force-velocity relation, the unusual tension-dependence of its Michaelis constant and the observed randomness of the kinesin motor (Visscher et al 1999;Thomas et al 2001). Furthermore, we discuss how neck-linker docking may lead to the occurrence of sub-8 nm steps (Coppin et al 1996;Nishiyama et al 2001). Figure 1a presents an idealized view of neck-linker docking (Rice et al 1999;Vale & Milligan 2000) for a single kinesin head attached to a β-tubulin monomer.…”

A modified active Brownian dynamics model using asymmetric energy conversion and its application to the molecular motor system

“…The coupling strength is reported to be weak when the rear head undertakes a biased diffusional search for its forward binding site. On the other hand, the coupling becomes strong when the new front head binds to the microtubule after releasing an ADP [18,47,48]. Hence the interaction strength is not a constant during a one-step cycle.…”

“…In spite of the extensive single-molecule studies, how kinesin functions to generate the force for discrete translation on a single protofilament of the microtubule is not well understood, leaving many questions on, for instance, substeps, backsteps, and fraction of power stroke and diffusion to each step [10,[18][19][20][21][22][23].…”