Measuring molecular rupture forces between single actin filaments and actin-binding proteins

Proc Natl Acad Sci U S A. 2008 Jul 8;105(27):9221-6. doi: 10.1073/pnas.0706124105. Epub 2008 Jun 30.

Abstract

Actin-binding proteins (ABPs) regulate the assembly of actin filaments (F-actin) into networks and bundles that provide the structural integrity of the cell. Two of these ABPs, filamin and alpha-actinin, have been extensively used to model the mechanical properties of actin networks grown in vitro; however, there is a lack in the understanding of how the molecular interactions between ABPs and F-actin regulate the dynamic properties of the cytoskeleton. Here, we present a native-like assay geometry to test the rupture force of a complex formed by an ABP linking two quasiparallel actin filaments. We readily demonstrate the adaptability of this assay by testing it with two different ABPs: filamin and alpha-actinin. For filamin/actin and alpha-actinin/actin, we measured similar rupture forces of 40-80 pN for loading rates between 4 and 50 pN/s. Both ABP unfolding and conformational transition events were observed, demonstrating that both are important and may be a significant mechanism for the temporal regulation of the mechanical properties of the actin cytoskeleton. With this modular, single-molecule assay, a wide range of ABP/actin interactions can be studied to better understand cytoskeletal and cell dynamics.

Publication types

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

MeSH terms

  • Actin Cytoskeleton / metabolism*
  • Actinin / metabolism
  • Animals
  • Biomechanical Phenomena
  • Contractile Proteins / metabolism
  • Filamins
  • Gelsolin / metabolism
  • Humans
  • Microfilament Proteins / metabolism*
  • Models, Molecular
  • Rabbits

Substances

  • Contractile Proteins
  • Filamins
  • Gelsolin
  • Microfilament Proteins
  • Actinin