Mechanism of regulation of Acanthamoeba myosin-IC by heavy-chain phosphorylation.
The ATPase activity of myosin-Is from lower eukaryotes is activated by phosphorylation by the p21-activated kinase family at the TEDS site on an actin-binding surface-loop. This actin-binding loop is the site of a cardiac myosin-II mutation responsible for some forms of familial hypertrophic cardiomyopathy. To determine the mechanism of myosin-I regulation by heavy-chain phosphorylation (HCP) and to better understand the importance of this loop in the function of all myosin isoforms, we performed a kinetic investigation of the regulatory mechanism of the Acanthamoeba myosin-IC motor domain (MIC(IQ)). Phosphorylated and dephosphorylated MIC(IQ) show actin-activated ATPase activity; however, HCP increases the ATPase activity >20-fold. HCP does not greatly affect the rate of phosphate release from MIC in the absence of actin, as determined by single turnover experiments. Additionally, HCP does not significantly affect the affinity of myosin for actin in the absence or presence of ATP, the rate of ATP-induced dissociation of actoMIC(IQ), the affinity of ADP, or the rate of ADP release. Sequential-mix single-turnover experiments show that HCP regulates the rate of phosphate release from actin-bound MIC(IQ). We propose that the TEDS-containing actin-binding loop plays a direct role in regulating phosphate release and the force-generating (A-to-R) transition of myosin-IC.
D000048:Acanthamoeba / Q000737:chemistry* / Q000201:enzymology / Q000235:genetics; D000199:Actins / Q000737:chemistry; D000244:Adenosine Diphosphate / Q000737:chemistry; D000251:Adenosine Triphosphatases / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D020816:Amino Acid Motifs; D000818:Animals; D005822:Genetic Vectors / Q000737:chemistry; D007700:Kinetics; D008956:Models, Chemical; D018995:Myosin Heavy Chains / Q000737:chemistry* / Q000235:genetics; D018994:Myosin Light Chains / Q000737:chemistry / Q000235:genetics; D024461:Myosin Type I / Q000737:chemistry* / Q000235:genetics; D028044:Phosphate-Binding Proteins / Q000737:chemistry; D010710:Phosphates / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D011817:Rabbits
D000199:Actins; D018994:Myosin Light Chains; D028044:Phosphate-Binding Proteins; D010710:Phosphates; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D024461:Myosin Type I; D018995:Myosin Heavy Chains