TY - JOUR
T1 - Different phosphorylated forms of myosin in contracting tracheal smooth muscle
AU - Persechini, A.
AU - Kamm, K. E.
AU - Stull, J. T.
N1 - Copyright:
Copyright 2004 Elsevier B.V., All rights reserved.
PY - 1986
Y1 - 1986
N2 - Calmodulin-dependent myosin light chain kinase phosphorylates two light chain subunits on each myosin molecule. We have developed a method for measuring nonphosphorylated, monophosphorylated, and diphosphorylated forms of myosin in smooth muscle. Four protein bands were separated in tissue extracts by nondenaturing polyacrylamide gel electrophoresis in the presence of pyrophosphate. Immunoblots demonstrated that three forms (designated M, MP, and MP2) reacted with rabbit antisera prepared against the purified phosphorylated light chain (P-light chain) from bovine tracheal smooth muscle. Evidence was obtained that M, MP, and MP2 represented nonphosphorylated, monophosphorylated, and diphosphorylated myosin, respectively, and that the other protein band was probably filamin. The formation of different phosphorylated forms of myosin was measured in bovine trachealis strips neurally stimulated from 1.0 to 3.5 s and quick-frozen. There was no detectable MP or MP2 in unstimulated muscles; the extent of P-light chain phosphorylation measured directly was 0.02 ± 0.01 mol of phosphate/mol of P-light chain. After 2.5-s stimulation, maximal values of 0.63 ± 0.06 mol of phosphate/mol of P-light chain and 0.40 ± 0.06 MP2/myosin(total) were obtained. During continuous neural stimulation from 1.0 to 3.5 s, the relationship between the extent of P-light chain phoshorylation (measured directly or calculated) and the relative amount of MP2 is consistent with a random phosphorylation process.
AB - Calmodulin-dependent myosin light chain kinase phosphorylates two light chain subunits on each myosin molecule. We have developed a method for measuring nonphosphorylated, monophosphorylated, and diphosphorylated forms of myosin in smooth muscle. Four protein bands were separated in tissue extracts by nondenaturing polyacrylamide gel electrophoresis in the presence of pyrophosphate. Immunoblots demonstrated that three forms (designated M, MP, and MP2) reacted with rabbit antisera prepared against the purified phosphorylated light chain (P-light chain) from bovine tracheal smooth muscle. Evidence was obtained that M, MP, and MP2 represented nonphosphorylated, monophosphorylated, and diphosphorylated myosin, respectively, and that the other protein band was probably filamin. The formation of different phosphorylated forms of myosin was measured in bovine trachealis strips neurally stimulated from 1.0 to 3.5 s and quick-frozen. There was no detectable MP or MP2 in unstimulated muscles; the extent of P-light chain phosphorylation measured directly was 0.02 ± 0.01 mol of phosphate/mol of P-light chain. After 2.5-s stimulation, maximal values of 0.63 ± 0.06 mol of phosphate/mol of P-light chain and 0.40 ± 0.06 MP2/myosin(total) were obtained. During continuous neural stimulation from 1.0 to 3.5 s, the relationship between the extent of P-light chain phoshorylation (measured directly or calculated) and the relative amount of MP2 is consistent with a random phosphorylation process.
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M3 - Article
C2 - 3516992
AN - SCOPUS:0023019582
SN - 0021-9258
VL - 261
SP - 6293
EP - 6299
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 14
ER -