Role of sequence 18-29 on actin in actomyosin interactions.
Affinity-purified polyclonal antibodies prepared against a synthetic peptide corresponding to sequence 18-29 from the N-terminus of rabbit alpha-skeletal actin reacted with G- and F-actin. Epitope mapping experiments with thrombin and hydroxylamine cleaved actin, and immunochemical assays verified the specificity of antibodies for the 18-29 sequence on actin. The binding of up to 0.5 mol of IgG per mole of actin did not affect the rigor binding of myosin subfragment 1 (S-1) to actin. Similarly, the binding of IgG to actin was not changed by a complete saturation of actin by S-1. In contrast to this, the weak acto-S-1 interactions in the presence of ATP were strongly inhibited by the 18-29 antibodies. At 25 degrees C, the acto-S-1 ATPase activity was inhibited by IgG stronger than the binding of S-1.ATP gamma S to actin. Thus, at this temperature, a catalytic inhibition of the acto-S-1 system appears to account at least in part for the antibody effect. Acto-S-1 ATPase activities at 25 degrees C were inhibited also by F(ab)(18-29). At 5 degrees C, the acto-S-1 ATPase activity and the binding of S-1.ATP to actin were inhibited approximately to the same extent by IgG(18-29). These results are discussed in terms of S-1 binding sites on actin and the possible role of sequence 18-29 in actomyosin interactions.
Adams
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S
Department of Chemistry and Biochemistry, University of California, Los Angeles 90024.
Reisler
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E
D000199:Actins / Q000276:immunology / Q000378:metabolism*; D000205:Actomyosin / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D000906:Antibodies / Q000276:immunology; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D007074:Immunoglobulin G / Q000378:metabolism; D015879:Myosin Subfragments / Q000378:metabolism; D010446:Peptide Fragments / Q000276:immunology / Q000378:metabolism*; D011817:Rabbits; D013917:Thrombin / Q000378:metabolism
D000199:Actins; D000906:Antibodies; D000939:Epitopes; D007074:Immunoglobulin G; D015879:Myosin Subfragments; D010446:Peptide Fragments; D000255:Adenosine Triphosphate; D000205:Actomyosin; D013917:Thrombin