Distinguishing inchworm and hand-over-hand processive kinesin movement by neck rotation measurements.
The motor enzyme kinesin makes hundreds of unidirectional 8-nanometer steps without detaching from or freely sliding along the microtubule on which it moves. We investigated the kinesin stepping mechanism by immobilizing a Drosophila kinesin derivative through the carboxyl-terminal end of the neck coiled-coil domain and measuring orientations of microtubules moved by single enzyme molecules at submicromolar adenosine triphosphate concentrations. The kinesin-mediated microtubule-surface linkage was sufficiently torsionally stiff (>/=2.0 +/- 0.9 x 10(-20) Newton meters per radian2) that stepping by the hypothesized symmetric hand-over-hand mechanism would produce 180 degree rotations of the microtubule relative to the immobilized kinesin neck. In fact, there were no rotations, a finding that is inconsistent with symmetric hand-over-hand movement. An alternative "inchworm" mechanism is consistent with our experimental results.
Hua
Wei
W
Biophysics and Structural Biology Program, Biochemistry Department, Brandeis University, Waltham, MA 02454-9110, USA.
Chung
Johnson
J
Gelles
Jeff
J
D000255:Adenosine Triphosphate / Q000378:metabolism; D000266:Adenylyl Imidodiphosphate / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D002384:Catalysis; D019281:Dimerization; D004330:Drosophila; D004800:Enzymes, Immobilized; D016547:Kinesins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D008870:Microtubules / Q000201:enzymology / Q000502:physiology*; D008954:Models, Biological; D020409:Molecular Motor Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D009068:Movement; D017434:Protein Structure, Tertiary; D012399:Rotation; D019809:Streptavidin
D004800:Enzymes, Immobilized; D020409:Molecular Motor Proteins; D000266:Adenylyl Imidodiphosphate; D000255:Adenosine Triphosphate; D019809:Streptavidin; D016547:Kinesins