title: Conformation-dependent ligand regulation of ATP hydrolysis by human KSP: activation of basal hydrolysis and inhibition of microtubule-stimulated hydrolysis by a single, small molecule modulator.
abstract: Human kinesin spindle protein (KSP)/hsEg5, a member of the kinesin-5 family, is essential for mitotic spindle assembly in dividing human cells and is required for cell cycle progression through mitosis. Inhibition of the ATPase activity of KSP leads to cell cycle arrest during mitosis and subsequent cell death. Ispinesib (SB-715992), a potent and selective inhibitor of KSP, is currently in phase II clinical trials for the treatment of multiple tumor types. Mutations that attenuate Ispinesib binding to KSP in vitro have been identified, highlighting the need for inhibitors that target different binding sites and inhibit KSP activity by novel mechanisms. We report here a small-molecule modulator, KSPA-1, that activates KSP-catalyzed ATP hydrolysis in the absence of microtubules yet inhibits microtubule-stimulated ATP hydrolysis by KSP. KSPA-1 inhibits cell proliferation and induces monopolar-spindle formation in tumor cells. Results from kinetic analyses, microtubule (MT) binding competition assays, and hydrogen/deuterium-exchange studies show that KSPA-1 does not compete directly for microtubule binding. Rather, this compound acts by driving a conformational change in the KSP motor domain and disrupts productive ATP turnover stimulated by MT. These findings provide a novel mechanism for targeting KSP and perhaps other mitotic kinesins.
authors.affiliation: Department of Enzymology and Mechanistic Pharmacology, GlaxoSmithKline, Collegeville, Pennsylvania 19426, USA. lusong.luo@gsk.com
authors.forename: Lusong
authors.initials: L
authors.lastname: Luo
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Carson
authors.forename: Kathleen S
authors.initials: KS
authors.lastname: Molnar
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Tuske
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Coales
authors.forename: Yoshitomo
authors.initials: Y
authors.lastname: Hamuro
authors.forename: Chiu-mei
authors.initials: CM
authors.lastname: Sung
authors.forename: Valery
authors.initials: V
authors.lastname: Sudakin
authors.forename: Kurt R
authors.initials: KR
authors.lastname: Auger
authors.forename: Dashyant
authors.initials: D
authors.lastname: Dhanak
authors.forename: Jeffrey R
authors.initials: JR
authors.lastname: Jackson
authors.forename: Pearl S
authors.initials: PS
authors.lastname: Huang
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Tummino
authors.forename: Robert A
authors.initials: RA
authors.lastname: Copeland
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001667:Binding, Competitive; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D003903:Deuterium / Q000378:metabolism; D006801:Humans; D006845:Hydrocarbons, Fluorinated / Q000494:pharmacology*; D006859:Hydrogen / Q000378:metabolism; D006868:Hydrolysis / Q000187:drug effects; D016547:Kinesins / Q000037:antagonists & inhibitors / Q000187:drug effects* / Q000378:metabolism; D008024:Ligands; D008298:Maleates / Q000494:pharmacology; D008870:Microtubules / Q000187:drug effects* / Q000378:metabolism; D011758:Pyrroles / Q000494:pharmacology*; D008941:Spindle Apparatus / Q000187:drug effects
chemical_list: D006845:Hydrocarbons, Fluorinated; C470243:KIF11 protein, human; C530009:KSPA-1; D008024:Ligands; D008298:Maleates; D011758:Pyrroles; D000244:Adenosine Diphosphate; D006859:Hydrogen; D000255:Adenosine Triphosphate; D003903:Deuterium; D016547:Kinesins