ADP-induced rocking of the kinesin motor domain revealed by single-molecule fluorescence polarization microscopy.
Kinesin is an ATP-driven molecular motor protein that moves processively along microtubules. Despite considerable research, the detailed mechanism of kinesin motion remains elusive. We applied an enhanced suite of single- and multiple-molecule fluorescence polarization microscopy assays to report the orientation and mobility of kinesin molecules bound to microtubules as a function of nucleotide state. In the presence of analogs of ATP, ADP-Pi or in the absence of nucleotide, the kinesin head maintains a rigid orientation. In the presence of ADP, the motor domain of kinesin, still bound to the microtubule, adopts a previously undescribed, highly mobile state. This state may be general to the chemomechanical cycle of motor proteins; in the case of kinesin, the transition from a highly mobile to a rigid state after ADP release may contribute to the generation of the 8 nm step.
Sosa
H
H
Department of Cellular and Molecular Medicine and Howard Hughes Medical Institute, University of California San Diego, La Jolla, California 92093-0683, USA. hsosa@aecom.yu.edu
Peterman
E J
EJ
Moerner
W E
WE
Goldstein
L S
LS
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000266:Adenylyl Imidodiphosphate / Q000378:metabolism; D017607:Aluminum Compounds / Q000378:metabolism; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes; D005459:Fluorides / Q000378:metabolism; D006801:Humans; D016547:Kinesins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D020409:Molecular Motor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D009068:Movement; D018583:Pliability; D011485:Protein Binding; D015202:Protein Engineering; D017434:Protein Structure, Tertiary
D017607:Aluminum Compounds; D005456:Fluorescent Dyes; D020409:Molecular Motor Proteins; C050992:tetrafluoroaluminate; D000266:Adenylyl Imidodiphosphate; D000244:Adenosine Diphosphate; D016547:Kinesins; D005459:Fluorides