title: Kinesin-1 structural organization and conformational changes revealed by FRET stoichiometry in live cells.
abstract: Kinesin motor proteins drive the transport of cellular cargoes along microtubule tracks. How motor protein activity is controlled in cells is unresolved, but it is likely coupled to changes in protein conformation and cargo association. By applying the quantitative method fluorescence resonance energy transfer (FRET) stoichiometry to fluorescent protein (FP)-labeled kinesin heavy chain (KHC) and kinesin light chain (KLC) subunits in live cells, we studied the overall structural organization and conformation of Kinesin-1 in the active and inactive states. Inactive Kinesin-1 molecules are folded and autoinhibited such that the KHC tail blocks the initial interaction of the KHC motor with the microtubule. In addition, in the inactive state, the KHC motor domains are pushed apart by the KLC subunit. Thus, FRET stoichiometry reveals conformational changes of a protein complex in live cells. For Kinesin-1, activation requires a global conformational change that separates the KHC motor and tail domains and a local conformational change that moves the KHC motor domains closer together.
authors.affiliation: Biophysics Research Division, University of Michigan, Ann Arbor, MI 48109, USA.
authors.forename: Dawen
authors.initials: D
authors.lastname: Cai
authors.forename: Adam D
authors.initials: AD
authors.lastname: Hoppe
authors.forename: Joel A
authors.initials: JA
authors.lastname: Swanson
authors.forename: Kristen J
authors.initials: KJ
authors.lastname: Verhey
mesh_terms: D000818:Animals; D019556:COS Cells; D002470:Cell Survival; D002522:Chlorocebus aethiops; D004789:Enzyme Activation; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D016547:Kinesins / Q000737:chemistry*; D008869:Microtubule-Associated Proteins / Q000737:chemistry / Q000378:metabolism; D008954:Models, Biological; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D021381:Protein Transport; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism
chemical_list: D008869:Microtubule-Associated Proteins; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D016547:Kinesins