Effect of α-actinin on actin structure Release of bound nucleotide

Abstract

We have examined the α-actinin-F-actin interaction by measuring the effect of highly purified α-actinin on bound nucleotide exchange in F-actin. Exchange was followed by measuring the release of actin-bound [¹⁴C]ADP in the presence of ATP using an ultrafiltration technique. α-Actinin increases by about 60 to 70% the rate of release of F-actin bound nucleotide when incubated for 1 h in the presence of 1 mM ATP/1 mM MgCl₂/0.05 mM CaCl₂/0.5 mM dithioerythritol/100 mM KCl/20 mM Tris-acetate, pH 7.5, at 37°C. The ability of α-actinin to enhance nucleotide exchange was maximal when α-actinin was added at a level near 10% of actin present by weight (molar ratio of 1 α-actinin to 49 actin monomers). The potentiating effect of α-actinin on the nucleotide exchange rate of F-actin was not highly related to the Mg²⁺: ATP ratio present in the incubation mixture. α-Actinin also increased the rate of bound nucleotide exchange of F-actin when the actin was present in a reconstituted actomyosin suspension. The results are consistent with the possibility that one α-actinin can affect the structure of multiple actin monomers present in an actin filament.