TY  - JOUR
A1  - Kelly, Alexander E.
A1  - Kranitz, Heather
A1  - Dötsch, Volker
A1  - Mullins, R. Dyche
T1  - Actin binding to the central domain of WASP/Scar proteins plays a critical role in the activation of the Arp2/3 complex
T2  - Journal of biological chemistry
N2  - 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.
Y1  - 2021
UR  - http://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/76250
UR  - https://nbn-resolving.org/urn:nbn:de:hebis:30:3-762504
SN  - 0021-9258
VL  - 281.2006
IS  - 15
SP  - 10589
EP  - 10597
PB  - American Society for Biochemistry and Molecular Biology Publications
CY  - Bethesda, Md
ER  -