Nucleotide-induced changes in the interaction of myosin subfragment 1 with actin: detection by antibodies against the N-terminal segment of actin.
The binding of myosin subfragment 1 (S-1) to actin in the presence and absence of nucleotides was determined under conditions of partial saturation of actin, up to 80%, by Fab(1-7), the antibodies against the first seven N-terminal residues on actin. In the absence of nucleotides, the binding constant of S-1 to actin (2 x 10(7) M-1) was decreased by 1 order of magnitude by Fab(1-7). The binding of S-1 to actin caused only limited displacement of Fab, and between 30 and 50% of actin appeared to bind both proteins. In the presence of MgAMP.PNP, MgADP, and MgPPi and at low S-1 concentrations, the same antibodies caused a large decrease in the binding of S-1 to actin. However, the binding of S-1.nucleotide to actin in the presence of Fab(1-7) increased cooperatively with the increase in S-1 concentration. Also, in contrast to rigor conditions, there was no indication for the binding of Fab(1-7) and S-1.nucleotide to the same actin molecules. These results show a nucleotide-induced transition in the actomyosin interface, most likely related to the different roles of the N-terminal segment of actin in the binding of S-1 and S-1.nucleotide. The possible implications of these findings to the regulation of actomyosin interactions are discussed.
DasGupta
G
G
Department of Chemistry and Biochemistry, University of California, Los Angeles 90024.
Reisler
E
E
D000199:Actins / Q000737:chemistry* / Q000187:drug effects / Q000276:immunology; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D001665:Binding Sites; D015879:Myosin Subfragments / Q000737:chemistry* / Q000187:drug effects / Q000276:immunology; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology*; D011685:Purine Nucleotides / Q000494:pharmacology*; D011817:Rabbits; D013329:Structure-Activity Relationship
D000199:Actins; D000906:Antibodies; D015879:Myosin Subfragments; D010446:Peptide Fragments; D011685:Purine Nucleotides