Correlation Between Self-Association Modes and GTPase Activation of Dynamin (original) (raw)

Abstract

The GTPase activity of dynamin is obligatorily coupled, by a mechanism yet unknown, to the internalization of clathrin-coated endocytic vesicles. Dynamin oligomerizes in vitro and in vivo and both its mechanical and enzymatic activities appear to be mediated by this self-assembly. In this study we demonstrate that dynamin is characterized by a tetramer/monomer equilibrium with an equilibrium constant of 1.67 × 1017 M−3. Stopped-flow fluorescence experiments show that the association rate constant for 2′(3′)-O-N-methylanthraniloyl (mant)GTP is 7.0 × 10−5 M−1 s−1 and the dissociation rate constant is 2.1 s−1, whereas the dissociation rate constant for mantdeoxyGDP is 93 s−1. We also demonstrate the cooperativity of dynamin binding and GTPase activation on a microtubule lattice. Our results indicate that dynamin self-association is not a sufficient condition for the expression of maximal GTPase activity, which suggests that dynamin molecules must be in the proper conformation or orientation if they are to form an active oligomer.

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Authors and Affiliations

  1. Department of Pharmacology, U.T. Southwestern Medical Center, Dallas, Texas, 75235–9041
    Derk D. Binns, Barbara Barylko, Nikolai Grichine & Joseph P. Albanesi
  2. Department of Biochemistry, U.T. Health Science Center, Tyler, Texas
    Mark A. L. Atkinson
  3. Department of Genetics and Molecular Biology, University of Hawaii, Honolulu, Hawaii, 96822
    Michael K. Helms & David M. Jameson
  4. Division of Physical Biochemistry, National Institute for Medical Research, Mill Hill, London, NW7 1AA, United Kingdom
    John F. Eccleston

Authors

  1. Derk D. Binns
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  2. Barbara Barylko
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  3. Nikolai Grichine
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  4. Mark A. L. Atkinson
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  5. Michael K. Helms
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  6. David M. Jameson
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  7. John F. Eccleston
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  8. Joseph P. Albanesi
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Binns, D.D., Barylko, B., Grichine, N. et al. Correlation Between Self-Association Modes and GTPase Activation of Dynamin.J Protein Chem 18, 277–290 (1999). https://doi.org/10.1023/A:1021083211267

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