X-ray scattering study of actin polymerization nuclei assembled by tandem W domains

Proc Natl Acad Sci U S A. 2008 Aug 5;105(31):10785-90. doi: 10.1073/pnas.0801650105. Epub 2008 Jul 31.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Actins / chemistry*
  • Actins / genetics
  • Amino Acid Sequence
  • Models, Molecular*
  • Molecular Sequence Data
  • Polymers / chemistry*
  • Protein Engineering / methods
  • Protein Structure, Tertiary
  • Wiskott-Aldrich Syndrome Protein Family / chemistry
  • X-Ray Diffraction

Substances

  • Actins
  • Polymers
  • Wiskott-Aldrich Syndrome Protein Family