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Actin binding to the central domain of WASP/Scar proteins plays a critical role in the activation of the Arp2/3 complex
- The Arp2/3 complex nucleates and cross-links actin filaments at the leading edge of motile cells, and its activity is stimulated by C-terminal regions of WASP/Scar proteins, called VCA domains. VCA domains contain a verprolin homology sequence (V) that binds monomeric actin and central (C) and acidic sequences (A) that bind the Arp2/3 complex. Here we show that the C domain binds to monomeric actin with higher affinity (K(d) = 10 microm) than to the Arp2/3 complex (K(d) > 200 microm). Nuclear magnetic resonance spectroscopy reveals that actin binds to the N-terminal half of the C domain and that both the V and C domains can bind actin independently and simultaneously, indicating that they interact with different sites. Mutation of conserved hydrophobic residues in the actin-binding interface of the C domain disrupts activation of the Arp2/3 complex but does not alter affinity for the complex. By chemical cross-linking the C domain interacts with the p40 subunit of the Arp2/3 complex and, by fluorescence polarization anisotropy, the binding of actin and the Arp2/3 complex are mutually exclusive. Our results indicate that both actin and Arp2/3 binding are important for C domain function but that the C domain does not form a static bridge between the two. We propose a model for activation of the Arp2/3 complex in which the C domain first primes the complex by inducing a necessary conformational change and then initiates nucleus assembly by bringing an actin monomer into proximity of the primed complex.
Author: | Alexander E. KellyORCiD, Heather Kranitz, Volker DötschORCiDGND, R. Dyche MullinsORCiD |
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URN: | urn:nbn:de:hebis:30:3-762504 |
DOI: | https://doi.org/10.1074/jbc.M507470200 |
ISSN: | 0021-9258 |
Pubmed Id: | https://pubmed.ncbi.nlm.nih.gov/16403731 |
Parent Title (English): | Journal of biological chemistry |
Publisher: | American Society for Biochemistry and Molecular Biology Publications |
Place of publication: | Bethesda, Md |
Document Type: | Article |
Language: | English |
Date of Publication (online): | 2021/01/04 |
Year of first Publication: | 2006 |
Publishing Institution: | Universitätsbibliothek Johann Christian Senckenberg |
Release Date: | 2024/04/22 |
Volume: | 281.2006 |
Issue: | 15 |
Page Number: | 9 |
First Page: | 10589 |
Last Page: | 10597 |
Institutes: | Biochemie, Chemie und Pharmazie / Biochemie und Chemie |
Dewey Decimal Classification: | 5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie |
Sammlungen: | Universitätspublikationen |
Licence (German): | Creative Commons - CC BY - Namensnennung 4.0 International |