title: Working stroke of the kinesin-14, ncd, comprises two substeps of different direction.
abstract: Single-molecule experiments have been used with great success to explore the mechanochemical cycles of processive motor proteins such as kinesin-1, but it has proven difficult to apply these approaches to nonprocessive motors. Therefore, the mechanochemical cycle of kinesin-14 (ncd) is still under debate. Here, we use the readout from the collective activity of multiple motors to derive information about the mechanochemical cycle of individual ncd motors. In gliding motility assays we performed 3D imaging based on fluorescence interference contrast microscopy combined with nanometer tracking to simultaneously study the translation and rotation of microtubules. Microtubules gliding on ncd-coated surfaces rotated around their longitudinal axes in an [ATP]- and [ADP]-dependent manner. Combined with a simple mechanical model, these observations suggest that the working stroke of ncd consists of an initial small movement of its stalk in a lateral direction when ADP is released and a second, main component of the working stroke, in a longitudinal direction upon ATP binding.
authors.affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany; Center for Molecular Bioengineering (B CUBE), Technische Universität Dresden, 01069 Dresden, Germany.
authors.forename: Bert
authors.initials: B
authors.lastname: Nitzsche
authors.affiliation: Nencki Institute of Experimental Biology, 02-093 Warsaw, Poland.
authors.forename: Elzbieta
authors.initials: E
authors.lastname: Dudek
authors.affiliation: Nencki Institute of Experimental Biology, 02-093 Warsaw, Poland.
authors.forename: Lukasz
authors.initials: L
authors.lastname: Hajdo
authors.affiliation: Nencki Institute of Experimental Biology, 02-093 Warsaw, Poland.
authors.forename: Andrzej A
authors.initials: AA
authors.lastname: Kasprzak
authors.affiliation: J. Stefan Institute, 1000 Ljubljana, Slovenia andrej.vilfan@ijs.si diez@bcube-dresden.de.
authors.forename: Andrej
authors.identifier: 0000-0001-8985-6072
authors.initials: A
authors.lastname: Vilfan
authors.affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany; Center for Molecular Bioengineering (B CUBE), Technische Universität Dresden, 01069 Dresden, Germany; andrej.vilfan@ijs.si diez@bcube-dresden.de.
authors.forename: Stefan
authors.initials: S
authors.lastname: Diez
mesh_terms: D000244:Adenosine Diphosphate / Q000737:chemistry* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000737:chemistry* / Q000378:metabolism; D001681:Biological Assay; D001696:Biomechanical Phenomena; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D016547:Kinesins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008870:Microtubules / Q000737:chemistry* / Q000648:ultrastructure; D008956:Models, Chemical; D015513:Oncogene Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012399:Rotation
chemical_list: D015513:Oncogene Proteins; D011993:Recombinant Fusion Proteins; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; C502598:KIF14 protein, human; D016547:Kinesins
keywords: fluorescence interference contrast microscopy; gliding motility assay; kinesin; mathematical modeling; microtubule