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    Differentiating between models of Epothilone binding to microtubules using tubulin mutagenesis, cytotoxicity, and molecular modeling

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    Entwistle_Wiley_2012.pdf (1004.Kb)
    Issue Date
    2012-09
    Author
    Entwistle, Ruth A.
    Rizk, Rania S.
    Cheng, Daniel M.
    Lushington, Gerald H.
    Himes, Richard H.
    Gupta, Mohan L., Jr
    Publisher
    Wiley
    Type
    Article
    Article Version
    Scholarly/refereed, author accepted manuscript
    Rights
    © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
    Metadata
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    Abstract
    Microtubule stabilizers are powerful anti-mitotic compounds and represent a proven cancer treatment strategy. Several classes of compounds in clinical use or trials, such as the taxanes and epothilones, bind to the same region of β-tubulin. Determining how these molecules interact with tubulin and stabilize microtubules is important both for understanding the mechanism of action and enhancing chemotherapeutic potential, e.g. reducing side effects, increasing solubility, and overcoming resistance. Structural studies using nonpolymerized tubulin or stabilized polymers have produced different models of epothilone binding. Here, we used directed mutagenesis of the binding site on Saccharomyces cerevisiae β-tubulin to analyze interactions between Epothilone B and its biologically relevant substrate, dynamic microtubules. Five engineered amino acid changes contributed to a 125-fold increase in Epothilone B cytotoxicity independent of inherent microtubule stability. The mutagenesis of endogenous β-tubulin was done in otherwise isogenic strains. This facilitated the correlation of amino acid substitutions with altered cytotoxicity using molecular mechanics simulations. The results, which are based on the interaction between Epothilone B and dynamic microtubules, most strongly support the binding mode determined by NMR spectroscopy-based studies. This work establishes a system for discriminating between potential binding modes and among various compounds and/or analogues using a sensitive biological activity-based readout.
    Description
    This is the peer reviewed version of the following article: Entwistle, R. A., Rizk, R. S., Cheng, D. M., Lushington, G. H., Himes, R. H., & Gupta, M. L. (2012). Differentiating between models of Epothilone binding to microtubules using tubulin mutagenesis, cytotoxicity, and molecular modeling. ChemMedChem, 7(9), 1580–1586. http://doi.org/10.1002/cmdc.201200286, which has been published in final form at doi.org/10.1002/cmdc.201200286. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.
    URI
    http://hdl.handle.net/1808/24017
    DOI
    https://doi.org/10.1002/cmdc.201200286
    Collections
    • Molecular Biosciences Scholarly Works [472]
    Citation
    Entwistle, R. A., Rizk, R. S., Cheng, D. M., Lushington, G. H., Himes, R. H., & Gupta, M. L. (2012). Differentiating between models of Epothilone binding to microtubules using tubulin mutagenesis, cytotoxicity, and molecular modeling. ChemMedChem, 7(9), 1580–1586. http://doi.org/10.1002/cmdc.201200286

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    Contact KU ScholarWorks
    785-864-8983
    KU Libraries
    1425 Jayhawk Blvd
    Lawrence, KS 66045
    785-864-8983

    KU Libraries
    1425 Jayhawk Blvd
    Lawrence, KS 66045
    Image Credits
     

     

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