title: Monoclonal antibody assessment of tissue- and species-specific myosin light chain kinase isozymes.
abstract: Monoclonal antibodies raised against chicken gizzard smooth muscle myosin light chain kinase were used for immunological and structural studies of this enzyme. Epitope mapping of trypsin-digested chicken gizzard enzyme showed that MM-1, 2, 3, 4, 5, 6, and 7 bind to 65 kDa (trypsin-digested) and 60 kDa (chymotrypsin-digested) fragments which contain the catalytic domain of the kinase. Kinetic analysis demonstrated that MM-7 inhibited kinase activity competitively with respect to ATP and noncompetitively with respect to myosin light chain, thereby indicating that MM-7 binds at or near the ATP binding site of the enzyme. Immunoblot analysis revealed that all these antibodies (MM-1 to 12) reacted with the enzyme (130 kDa) from intestinal and vascular smooth muscles, whereas 5 (MM-1, 3, 4, 6, and 9) or 3 (MM-1, 3, and 4) of 12 antibodies did not cross-react with chicken cardiac muscle or with blood platelet myosin light chain kinase (130 kDa), respectively. None of these antibodies showed cross-reactivity against skeletal muscle myosin light chain kinase. As for mammalian species, MM-11 and 12 reacted with myosin light chain kinase of vascular smooth muscle (140 kDa) and MM-11 cross-reacted with the enzyme (140 kDa) from cardiac muscle of rat and rabbit. These data suggest the existence of at least 4 subspecies of myosin light chain kinase in chicken tissues and the heterogeneity of tissue- and species-specific isozyme forms.
authors: Department of Pharmacology, Nagoya University School of Medicine, Aichi.
authors: M
authors: M
authors: Hagiwara
authors: H
authors: H
authors: Tokumitsu
authors: K
authors: K
authors: Onoda
authors: T
authors: T
authors: Tanaka
authors: M
authors: M
authors: Ito
authors: N
authors: N
authors: Kato
authors: H
authors: H
authors: Hidaka
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D002645:Chickens; D000939:Epitopes / Q000032:analysis; D005895:Gizzard, Avian / Q000201:enzymology; D015151:Immunoblotting; D007527:Isoenzymes / Q000032:analysis / Q000276:immunology*; D007700:Kinetics; D009130:Muscle, Smooth / Q000201:enzymology; D009219:Myosin-Light-Chain Kinase / Q000032:analysis / Q000276:immunology*; D013045:Species Specificity; D014018:Tissue Distribution
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D007527:Isoenzymes; D009219:Myosin-Light-Chain Kinase