title: X-ray scattering study of actin polymerization nuclei assembled by tandem W domains.
abstract: The initiation of actin polymerization in cells requires actin filament nucleators. With the exception of formins, known filament nucleators use the Wiskott-Aldrich syndrome protein (WASP) homology 2 (WH2 or W) domain for interaction with actin. A common architecture, found in Spire, Cobl, VopL, and VopF, consists of tandem W domains that tie together three to four actin monomers to form a polymerization nucleus. Uncontrollable polymerization has prevented the structural investigation of such nuclei. We have engineered stable nuclei consisting of an actin dimer and a trimer stabilized by tandem W domain hybrid constructs and studied their structures in solution by x-ray scattering. We show that Spire-like tandem W domains stabilize a polymerization nucleus by lining up actin subunits along the long-pitch helix of the actin filament. Intersubunit contacts in the polymerization nucleus, thought to involve the DNase I-binding loop of actin, coexist with the binding of the W domain in the cleft between actin subdomains 1 and 3. The successful stabilization of filament-like multiactin assemblies opens the way to the crystallographic investigation of intersubunit contacts in the actin filament.
authors.affiliation: Department of Physiology, School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors.forename: Grzegorz
authors.initials: G
authors.lastname: Rebowski
authors.forename: Malgorzata
authors.initials: M
authors.lastname: Boczkowska
authors.forename: David B
authors.initials: DB
authors.lastname: Hayes
authors.forename: Liang
authors.initials: L
authors.lastname: Guo
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Irving
authors.forename: Roberto
authors.initials: R
authors.lastname: Dominguez
mesh_terms: D000199:Actins / Q000737:chemistry* / Q000235:genetics; D000595:Amino Acid Sequence; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011108:Polymers / Q000737:chemistry*; D015202:Protein Engineering / Q000379:methods; D017434:Protein Structure, Tertiary; D051300:Wiskott-Aldrich Syndrome Protein Family / Q000737:chemistry; D014961:X-Ray Diffraction
chemical_list: D000199:Actins; D011108:Polymers; D051300:Wiskott-Aldrich Syndrome Protein Family