Identification of amino acid substitutions differentiating actin isoforms in their interaction with myosin.
Various aspects of actin--myosin interaction were studied with actin preparations from two types of smooth muscle: bovine aorta and chicken gizzard, and from two types of sarcomeric muscle: bovine cardiac and rabbit skeletal. All four preparations activated the Mg2+-ATPase activity of skeletal muscle myosin to the same Vmax, but the Kapp for the smooth muscle preparations was higher. At low KCl, pH 8.0 and millimolar substrate concentrations the Kapp values differed by a factor of 2.5. This differential behaviour of the four actin preparations correlates with amino acid substitutions at positions 17 and 89 of actin polypeptide chain, differentiating the smooth-muscle-specific gamma and alpha isomers from cardiac and skeletal-muscle-specific alpha isomers. This correlation provides evidence for involvement of the NH2-terminal portion of the actin polypeptide chain in the interaction with myosin. The differences in the activation of myosin ATPase by various actins were sensitive to changes in the substrate and KCl concentration and pH of the assay medium. Addition of myosin subfragment-1 or heavy meromyosin in the absence of nucleotide produced similar changes in the fluorescence of a fluorescent reagent N-(1-pyrenyl)-iodoacetamide, attached at Cys-374, or 1,N6-ethenoadenosine 5'-diphosphate substituted for the bound ADP in actin protomers in gizzard and skeletal muscle F-actin. The results are consistent with an influence of the amino acid substitutions on ionic interactions leading to complex formation between actin and myosin intermediates in the ATPase cycle but not on the associated states.
D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001011:Aorta / Q000378:metabolism; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism*; D002417:Cattle; D002645:Chickens; D004789:Enzyme Activation; D005895:Gizzard, Avian / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D009132:Muscles / Q000378:metabolism; D009206:Myocardium / Q000378:metabolism; D015879:Myosin Subfragments / Q000378:metabolism; D009218:Myosins / Q000378:metabolism*; D009711:Nucleotides / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D013045:Species Specificity; D013050:Spectrometry, Fluorescence; D013379:Substrate Specificity
D000199:Actins; D015879:Myosin Subfragments; D009711:Nucleotides; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins