Interaction between Myosin and F-Actin. Correlation with Actin-Binding Sites on Subfragment-11

Abstract

F-Actin bindings to subfragment-1 (S-1) and S-1 after limited proteolysis by trypsin (S-1_t) were studied in the absence and presence of ATP by means of ultracentrifugation. No significant difference in the affinities for F-actin was observed between S-1 and S-1_t in the absence of ATP. In contrast, the affinity for F-actin in the presence of ATP was decreased about 50 times by the limited proteolysis of the S-1 heavy chain. The S-1 whose SH₁ and SH₂ groups were cross-linked by N,N′-p-phenylenedimaleimide bound F-actin weakly. The affinity for F-actin was similar to that of unmodified S-1 in the presence of ATP and was also decreased markedly by limited proteolysis of the cross-linked S-1. Reciprocals of the dissociation constant of acto-S-1 complex decreased markedly with increase of ionic strength in the presence of ATP, but decreased only slightly at the rigor state. All these results are consistent with our proposal that S-1 has two different actin binding sites, as reported previously (Katoh, T., Imae, S., & Morita, F. (1984) J. Biochem, 95, 447–454). The mechanism of activation of S-1 ATPase by F-actin is discussed.