title: Measuring molecular rupture forces between single actin filaments and actin-binding proteins.
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.
authors.affiliation: Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Jorge M
authors.initials: JM
authors.lastname: Ferrer
authors.forename: Hyungsuk
authors.initials: H
authors.lastname: Lee
authors.forename: Jiong
authors.initials: J
authors.lastname: Chen
authors.forename: Benjamin
authors.initials: B
authors.lastname: Pelz
authors.forename: Fumihiko
authors.initials: F
authors.lastname: Nakamura
authors.forename: Roger D
authors.initials: RD
authors.lastname: Kamm
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Lang
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism*; D000185:Actinin / Q000378:metabolism; D000818:Animals; D001696:Biomechanical Phenomena; D003285:Contractile Proteins / Q000378:metabolism; D064448:Filamins; D018260:Gelsolin / Q000378:metabolism; D006801:Humans; D008840:Microfilament Proteins / Q000378:metabolism*; D008958:Models, Molecular; D011817:Rabbits
chemical_list: D003285:Contractile Proteins; D064448:Filamins; D018260:Gelsolin; D008840:Microfilament Proteins; D000185:Actinin