The Beginning of Kinesin's Force-Generating Cycle Visualized at 9-Å Resolution

• Published in: The Journal of cell biology Vol. 177; no. 3; pp. 377 - 385
• Main Authors: Sindelar, Charles V., Downing, Kenneth H.
• Format: Journal Article
• Language: English
• Published: United States Rockefeller University Press 07.05.2007; The Rockefeller University Press
• Subjects: Active sites; Adenosine diphosphate; Adenosine Diphosphate - chemistry; Adenosine Diphosphate - metabolism; Animals; Biochemistry; Cellular biology; Cryoelectron Microscopy; Crystal structure; Crystals; Electron microscopes; Humans; Image Processing, Computer-Assisted; Image reconstruction; Kinesin - chemistry; Kinesin - metabolism; Microtubules; Microtubules - chemistry; Microtubules - metabolism; Microtubules - ultrastructure; Models, Molecular; Molecules; Motors; Multiprotein Complexes - chemistry; Multiprotein Complexes - metabolism; Multiprotein Complexes - ultrastructure; Nucleotides; Peptides; Protein Binding - physiology; Protein Structure, Secondary; Proteins; Switches
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.200612090

We have used cryo-electron microscopy of kinesin-decorated microtubules to resolve the structure of the motor protein kinesin's crucial nucleotide response elements, switch I and the switch II helix, in kinesin's poorly understood nucleotide-free state. Both of the switch elements undergo conformational change relative to the microtubule-free state. The changes in switch I suggest a role for it in "ejecting" adenosine diphosphate when kinesin initially binds to the microtubule. The switch II helix has an N-terminal extension, apparently stabilized by conserved microtubule contacts, implying a microtubule activation mechanism that could convey the state of the bound nucleotide to kinesin's putative force-delivering element (the "neck linker"). In deriving this structure, we have adapted an image-processing technique, single-particle reconstruction, for analyzing decorated microtubules. The resulting reconstruction visualizes the asymmetric seam present in native, 13-protofilament microtubules, and this method will provide an avenue to higher-resolution characterization of a variety of microtubule-binding proteins, as well as the microtubule itself.