Journal of Biological Chemistry
Volume 283, Issue 38, 19 September 2008, Pages 25839-25845
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Molecular Basis of Cell and Developmental Biology
Regulatory ATPase Sites of Cytoplasmic Dynein Affect Processivity and Force Generation*

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The heavy chain of cytoplasmic dynein contains four nucleotide-binding domains referred to as AAA1–AAA4, with the first domain (AAA1) being the main ATP hydrolytic site. Although previous studies have proposed regulatory roles for AAA3 and AAA4, the role of ATP hydrolysis at these sites remains elusive. Here, we have analyzed the single molecule motility properties of yeast cytoplasmic dynein mutants bearing mutations that prevent ATP hydrolysis at AAA3 or AAA4. Both mutants remain processive, but the AAA4 mutant exhibits a surprising increase in processivity due to its tighter affinity for microtubules. In addition to changes in motility characteristics, AAA3 and AAA4 mutants produce less maximal force than wild-type dynein. These results indicate that the nucleotide binding state at AAA3 and AAA4 can allosterically modulate microtubule binding affinity and affect dynein processivity and force production.

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*

This work was supported, in whole or in part, by National Institutes of Health Grant T32 GM07810 (to C. C.). This work was also supported by the Howard Hughes Medical Institute (to R. D. V.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. S1 and S2 and Movies S1–S3.

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Present address: Dept. of Cell Biology, Harvard Medical School, Boston, MA 02115.