title: A model system of coupled activity of co-immobilized creatine kinase and myosin.
abstract: Myosin and creatine kinase were co-immobilized onto Immunodyne films to mimic the behaviour of creatine kinase bound to the M-line of myofilaments. The Mg-ATPase activity of bound myosin was studied by a coupled enzymatic assay, which detects Mg-ADP in the bulk solution by means of pyruvate kinase and lactate dehydrogenase. The competition for Mg-ADP between pyruvate kinase and creatine kinase either free in solution or co-immobilized with myosin was studied at various creatine phosphate concentrations. Bound creatine kinase competed efficiently when present in very low amounts, corresponding to an activity ratio higher than 1:20,000 between creatine kinase and pyruvate kinase and a molar ratio higher than 1:1000 between creatine kinase and myosin. The Mg-ADP produced by myosin ATPase in the vicinity of the film did not diffuse into the bulk solution but, in the presence of creatine phosphate, was recycled into Mg-ATP by the neighbouring creatine kinase. The existence of an unstirred layer near the surface of the film is sufficient to explain the channeling of ADP (or ATP) between co-immobilized myosin and creatine kinase, without direct interaction or 'intimate coupling' between the enzymes. The problem now is to determine the importance of this kind of facilitated diffusion in the myofilaments in vivo.
authors.affiliation: Laboratoire de Biologie Physico-Chimique, Université Paris-Sud, France.
authors.forename: M
authors.initials: M
authors.lastname: Arrio-Dupont
authors.forename: J J
authors.initials: JJ
authors.lastname: Béchet
authors.forename: A
authors.initials: A
authors.lastname: d'Albis
mesh_terms: D000818:Animals; D001667:Binding, Competitive; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism; D003402:Creatine Kinase / Q000378:metabolism*; D004800:Enzymes, Immobilized / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D011770:Pyruvate Kinase / Q000378:metabolism; D011817:Rabbits
chemical_list: D004800:Enzymes, Immobilized; D011770:Pyruvate Kinase; D003402:Creatine Kinase; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins