Nucleotide-dependent angular change in kinesin motor domain bound to tubulin.
Kinesin is a 'motor' molecule, consisting of two head domains, an alpha-helical coiled coil rod, and a tail part that binds to its cargo. When expressed in a bacterial system, the head domain is functional, and can bind to microtubules with the stoichiometry of one head per tubulin dimer. Kinesin moves along microtubules by means of a cyclic process of nucleotide binding, hydrolysis and product release. We have used negative-stain electron microscopy and image analysis to study the structures of microtubules and tubulin sheets decorated with the motor domain (head) of kinesin in three states: in the presence of an unhydrolysable ATP analogue, 5'-adenylylimidodiphosphate (AMP-PNP); without nucleotides; and with adenosine 5'-diphosphate (ADP). A single kinesin head bound to a microtubule has a pear-shaped structure, with the broader end towards the 'plus' end of the microtubule under all conditions; the reverse motor, ncd, is similarly oriented. Three-dimensional maps reveal that kinesin heads have a spike that is assumed to form the attachment to the tail of a complete kinesin molecule. This spike is perpendicular to the microtubule axis in the presence of ADP, but points towards the plus end (approximately 45 degrees) in the presence of AMP-PNP or absence of nucleotides. Our results provide direct evidence for a conformational change of the kinesin motor domain during the ATPase cycle.
Hirose
K
K
MRC Laboratory of Molecular Biology, Cambridge, UK.
Lockhart
A
A
Cross
R A
RA
Amos
L A
LA
D000266:Adenylyl Imidodiphosphate / Q000378:metabolism; D000818:Animals; D004330:Drosophila; D006135:Gryllidae; D007091:Image Processing, Computer-Assisted; D016547:Kinesins / Q000378:metabolism / Q000648:ultrastructure*; D011485:Protein Binding; D011487:Protein Conformation; D051381:Rats; D012617:Sea Urchins; D014404:Tubulin / Q000378:metabolism / Q000648:ultrastructure*
D014404:Tubulin; D000266:Adenylyl Imidodiphosphate; D016547:Kinesins