title: A chimeric kinesin-1 head/kinesin-5 tail motor switches between diffusive and processive motility.
abstract: Homotetrameric kinesin-5 motors are essential for chromosome separation and assembly of the mitotic spindle. These kinesins bind between two microtubules (MTs) and slide them apart, toward the spindle poles. This process must be tightly regulated in mitosis. In in vitro assays, Eg5 moves diffusively on single MTs and switches to a directed mode between MTs. How allosteric communication between opposing motor domains works remains unclear, but kinesin-5 tail domains may be involved. Here we present a single-molecule fluorescence study of a tetrameric kinesin-1 head/kinesin-5 tail chimera, DK4mer. This motor exhibited fast processive motility on single MTs interrupted by pauses. Like Eg5, DK4mer diffused along MTs with ADP, and slid antiparallel MTs apart with ATP. In contrast to Eg5, diffusive and processive periods were clearly distinguishable. This allowed us to measure transition rates among states and for unbinding as a function of buffer ionic strength. These data, together with results from controls using tail-less dimers, indicate that there are two modes of interaction with MTs, separated by an energy barrier. This result suggests a scheme of motor regulation that involves switching between two bound states, possibly allosterically controlled by the opposing tetramer end. Such a scheme is likely to be relevant for the regulation of native kinesin-5 motors.
authors: Drittes Physikalisches Institut, Georg-August-Universität Göttingen, Göttingen, Germany.
authors: Christina
authors: C
authors: Thiede
authors: Stefan
authors: S
authors: Lakämper
authors: Alok D
authors: AD
authors: Wessel
authors: Stefanie
authors: S
authors: Kramer
authors: Christoph F
authors: CF
authors: Schmidt
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003432:Cross-Linking Reagents / Q000378:metabolism; D004058:Diffusion; D029721:Drosophila Proteins / Q000737:chemistry / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000378:metabolism; D005453:Fluorescence; D049452:Green Fluorescent Proteins / Q000378:metabolism; D016547:Kinesins / Q000737:chemistry / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D009994:Osmolar Concentration; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029867:Xenopus Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D003432:Cross-Linking Reagents; D029721:Drosophila Proteins; C088335:KIF11 protein, Xenopus; D050505:Mutant Proteins; D011994:Recombinant Proteins; D029867:Xenopus Proteins; D049452:Green Fluorescent Proteins; C515209:Khc protein, Drosophila; D016547:Kinesins