title: Direct binding of myosin II to phospholipid vesicles via tail regions and phosphorylation of the heavy chains by protein kinase C.
abstract: Recent cloning and sequencing studies suggest that heavy chains of all non-muscle myosins II have a protein kinase C (PKC) phosphorylation site within their tail regions. A fragment of human macrophage myosin heavy chain, encompassing its COOH-terminal 396 amino acids (MIIAF46), was expressed in Escherichia coli to provide a model system for study of PKC-mediated phosphorylation. PKC phosphorylated this fragment when phosphatidylserine (PS) liposomes were present, but not when liposomes made from PS/phosphatidylcholine (PC) were used. The reaction required Ca2+, but not other activators such as diacylglycerol (DG) or phosphatidylinositol 4,5-bisphosphate. Phosphorylation of MIIAF46 was not observed in the presence of micelles of PS or PS/DG. Similar results were obtained using native myosin II purified from bovine brain and chicken intestine brush border. Phosphorylation of light chains, in contrast, occurred even with PS/PC liposomes if DG was present. Addition of the PS and PS/DG liposomes significantly increased the turbidities at 340 nm of MIIAF46 and native myosin II, and the extent of increase depended upon the type of myosin used. Also, PS and PS/DG liposomes shifted the gel filtration elution positions of MIIAF46 and myosin II. In contrast, liposomes of PS/PC and PS/PC/DG gave only a slight increase in turbidity with all myosins and fragments and did not noticeably shift their gel filtration elution positions. These results suggest that myosins II bind to PS liposomes via the COOH-terminal regions of their heavy chains with affinities specific to each myosin isoform, that the binding is dependent upon the PS composition, and that PKC phosphorylates the PS-bound heavy chains.
authors.affiliation: Department of Pharmacology, Institute for Basic Research in Developmental Disabilities, Staten Island, New York 10134.
authors.forename: N
authors.initials: N
authors.lastname: Murakami
authors.forename: M
authors.initials: M
authors.lastname: Elzinga
authors.forename: S S
authors.initials: SS
authors.lastname: Singh
authors.forename: V P
authors.initials: VP
authors.lastname: Chauhan
mesh_terms: D001483:Base Sequence; D047390:Casein Kinase II; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D008081:Liposomes / Q000378:metabolism; D008264:Macrophages; D008969:Molecular Sequence Data; D009218:Myosins / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism
chemical_list: D008081:Liposomes; D010446:Peptide Fragments; D010743:Phospholipids; D011994:Recombinant Proteins; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D011493:Protein Kinase C; D009218:Myosins