Kinesin Kar3Cik1 ATPase pathway for microtubule cross-linking.
J Biol Chem
; 286(33): 29261-29272, 2011 Aug 19.
Article
em En
| MEDLINE
| ID: mdl-21680740
ABSTRACT
Kar3Cik1 is a Saccharomyces cerevisiae kinesin-14 that functions to shorten cytoplasmic microtubules (MTs) during yeast mating yet maintains mitotic spindle stability by cross-linking anti-parallel interpolar MTs. Kar3 contains both an ATP- and a MT-binding site, yet there is no evidence of a nucleotide-binding site in Cik1. Presteady-state and steady-state kinetic experiments were pursued to define the regulation of Kar3Cik1 interactions with the MT lattice expected during interpolar MT cross-linking. The results reveal that association of Kar3Cik1 with the MT occurs at 4.9 µM(-1) s(-1), followed by a 5-s(-1) structural transition that limits ADP release from the Kar3 head. Mant-ATP binding occurred at 2.1 µM(-1) s(-1), and the pulse-chase experiments revealed an ATP-promoted isomerization at 69 s(-1). ATP hydrolysis was observed as a rapid step at 26 s(-1) and was required for the Kar3Cik1 motor to detach from MT. The conformational change at 5 s(-1) that occurred after Kar3Cik1 MT association and prior to ADP release was hypothesized to be the rate-limiting step for steady-state ATP turnover. We propose a model in which Kar3Cik1 interacts with the MT lattice through an alternating cycle of Cik1 MT collision followed by Kar3 MT binding with head-head communication between Kar3 and Cik1 modulated by the Kar3 nucleotide state and intramolecular strain.
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01-internacional
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MEDLINE
Assunto principal:
Saccharomyces cerevisiae
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Adenosina Trifosfatases
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Proteínas de Saccharomyces cerevisiae
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Proteínas Associadas aos Microtúbulos
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Proteínas dos Microtúbulos
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Microtúbulos
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Modelos Biológicos
Idioma:
En
Revista:
J Biol Chem
Ano de publicação:
2011
Tipo de documento:
Article