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Biophys J. 2014 Nov 4;107(9):2204-13. doi: 10.1016/j.bpj.2014.09.019.

Kinesin-5 allosteric inhibitors uncouple the dynamics of nucleotide, microtubule, and neck-linker binding sites.

Author information

1
Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, Michigan.
2
Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, Michigan. Electronic address: bjgrant@umich.edu.

Abstract

Kinesin motor domains couple cycles of ATP hydrolysis to cycles of microtubule binding and conformational changes that result in directional force and movement on microtubules. The general principles of this mechanochemical coupling have been established; however, fundamental atomistic details of the underlying allosteric mechanisms remain unknown. This lack of knowledge hampers the development of new inhibitors and limits our understanding of how disease-associated mutations in distal sites can interfere with the fidelity of motor domain function. Here, we combine unbiased molecular-dynamics simulations, bioinformatics analysis, and mutational studies to elucidate the structural dynamic effects of nucleotide turnover and allosteric inhibition of the kinesin-5 motor. Multiple replica simulations of ATP-, ADP-, and inhibitor-bound states together with network analysis of correlated motions were used to create a dynamic protein structure network depicting the internal dynamic coordination of functional regions in each state. This analysis revealed the intervening residues involved in the dynamic coupling of nucleotide, microtubule, neck-linker, and inhibitor binding sites. The regions identified include the nucleotide binding switch regions, loop 5, loop 7, ?4-?5-loop 13, ?1, and ?4-?6-?7. Also evident were nucleotide- and inhibitor-dependent shifts in the dynamic coupling paths linking functional sites. In particular, inhibitor binding to the loop 5 region affected ?-sheet residues and ?1, leading to a dynamic decoupling of nucleotide, microtubule, and neck-linker binding sites. Additional analyses of point mutations, including P131 (loop 5), Q78/I79 (?1), E166 (loop 7), and K272/I273 (?7) G325/G326 (loop 13), support their predicted role in mediating the dynamic coupling of distal functional surfaces. Collectively, our results and approach, which we make freely available to the community, provide a framework for explaining how binding events and point mutations can alter dynamic couplings that are critical for kinesin motor domain function.

PMID:
25418105
PMCID:
PMC4223232
DOI:
10.1016/j.bpj.2014.09.019
[Indexed for MEDLINE]
Free PMC Article

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