title: Kinesin processivity is gated by phosphate release.
abstract: Kinesin-1 is a dimeric motor protein, central to intracellular transport, that steps hand-over-hand toward the microtubule (MT) plus-end, hydrolyzing one ATP molecule per step. Its remarkable processivity is critical for ferrying cargo within the cell: over 100 successive steps are taken, on average, before dissociation from the MT. Despite considerable work, it is not understood which features coordinate, or "gate," the mechanochemical cycles of the two motor heads. Here, we show that kinesin dissociation occurs subsequent to, or concomitant with, phosphate (P(i)) release following ATP hydrolysis. In optical trapping experiments, we found that increasing the steady-state population of the posthydrolysis ADP · P(i) state (by adding free P(i)) nearly doubled the kinesin run length, whereas reducing either the ATP binding rate or hydrolysis rate had no effect. The data suggest that, during processive movement, tethered-head binding occurs subsequent to hydrolysis, rather than immediately after ATP binding, as commonly suggested. The structural change driving motility, thought to be neck linker docking, is therefore completed only upon hydrolysis, and not ATP binding. Our results offer additional insights into gating mechanisms and suggest revisions to prevailing models of the kinesin reaction cycle.
authors.affiliation: Biophysics Program.
authors.forename: Bojan
authors.initials: B
authors.lastname: Milic
authors.affiliation: Departments of Physics.
authors.forename: Johan O L
authors.initials: JO
authors.lastname: Andreasson
authors.affiliation: Department of Biomedical Engineering, Pennsylvania State University, University Park, PA 16802.
authors.forename: William O
authors.initials: WO
authors.lastname: Hancock
authors.affiliation: Biology, and Applied Physics, Stanford University, Stanford, CA 94305; and sblock@stanford.edu.
authors.forename: Steven M
authors.initials: SM
authors.lastname: Block
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D055592:Biophysical Phenomena; D029721:Drosophila Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006868:Hydrolysis; D016547:Kinesins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008954:Models, Biological; D020409:Molecular Motor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D052898:Optical Tweezers; D010710:Phosphates / Q000378:metabolism; D020836:Protein Structure, Quaternary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D029721:Drosophila Proteins; D020409:Molecular Motor Proteins; D010710:Phosphates; D011993:Recombinant Fusion Proteins; D000255:Adenosine Triphosphate; C515209:Khc protein, Drosophila; D016547:Kinesins
keywords: mechanochemistry; molecular motors; optical tweezers; single molecule