Allosteric Drug Discrimination Is Coupled to Mechanochemical Changes in the Kinesin-5 Motor Core |
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Authors: | Elizabeth D. Kim Rebecca Buckley Sarah Learman Jessica Richard Courtney Parke David K. Worthylake Edward J. Wojcik Richard A. Walker Sunyoung Kim |
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Affiliation: | From the ‡Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center, New Orleans, Louisiana 70112 and ;the §Department of Biological Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061 |
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Abstract: | Essential in mitosis, the human Kinesin-5 protein is a target for >80 classes of allosteric compounds that bind to a surface-exposed site formed by the L5 loop. Not established is why there are differing efficacies in drug inhibition. Here we compare the ligand-bound states of two L5-directed inhibitors against 15 Kinesin-5 mutants by ATPase assays and IR spectroscopy. Biochemical kinetics uncovers functional differences between individual residues at the N or C termini of the L5 loop. Infrared evaluation of solution structures and multivariate analysis of the vibrational spectra reveal that mutation and/or ligand binding not only can remodel the allosteric binding surface but also can transmit long range effects. Changes in L5-localized 310 helix and disordered content, regardless of substitution or drug potency, are experimentally detected. Principal component analysis couples these local structural events to two types of rearrangements in β-sheet hydrogen bonding. These transformations in β-sheet contacts are correlated with inhibitory drug response and are corroborated by wild type Kinesin-5 crystal structures. Despite considerable evolutionary divergence, our data directly support a theorized conserved element for long distance mechanochemical coupling in kinesin, myosin, and F1-ATPase. These findings also suggest that these relatively rapid IR approaches can provide structural biomarkers for clinical determination of drug sensitivity and drug efficacy in nucleotide triphosphatases. |
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Keywords: | Allosteric Regulation ATPases Drug Action Drug Resistance Fourier Transform IR (FTIR) Kinesin Molecular Motors Protein Structure Multivariate Data Analysis |
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