Sites phosphorylated in myosin light chain in contracting smooth muscle

J. C. Colburn, C. H. Michnoff, L. C. Hsu, Clive A. Slaughter, K. E. Kamm, J. T. Stull

Research output: Contribution to journalArticle

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Abstract

Purified smooth muscle myosin light chain can be phosphorylated at multiple sites by myosin light chain kinase and protein kinase C. We have determined the sites phosphorylated on myosin light chain in intact bovine tracheal smooth muscle. Stimulation with 10 μM carbachol resulted in 66 ± 5% monophosphorylated and 11 ± 2% diphosphorylated myosin light chain after 1 min, and 47 ± 4% monophosphorylated and 5 ± 2% diphosphorylated myosin light chain after 30 min. Myosin heavy chain contained 0.06 ± 0.01 mol of phosphate/mol of protein which did not change with carbachol. At both 1 and 30 min the monophosphorylated myosin light chain contained only phosphoserine whereas the diphosphorylated myosin light chain contained both phosphoserine and phosphothreonine. Two-dimensional peptide mapping of tryptic digests of monophosphorylated and diphosphorylated myosin light chain obtained from carbachol-stimulated tissue was similar to the peptide maps of purified light chain monophosphorylated and diphosphorylated, respectively, by myosin light chain kinase; these maps were distinct from the map obtained with tracheal light chain phosphorylated by protein kinase C. Phosphorylation of tracheal smooth muscle myosin light chain by myosin light chain kinase yields the tryptic phosphopeptide ATSNVFAMFDQSQIQEFK with S the phosphoserine in the monophosphorylated myosin light chain and TS the phosphothreonine and phosphoserine in the diphosphorylated myosin light chain. Thus, stimulation of tracheal smooth muscle with a high concentration of carbachol results in formation of both monophosphorylated and diphosphorylated myosin light chain although the amount of diphosphorylated light chain is substantially less than monophosphorylated light chain. In the intact muscle, myosin light chain is phosphorylated at sites corresponding to myosin light chain kinase phosphorylation.

Original languageEnglish (US)
Pages (from-to)19166-19173
Number of pages8
JournalJournal of Biological Chemistry
Volume263
Issue number35
StatePublished - Dec 1 1988
Externally publishedYes

Fingerprint

Myosin Light Chains
Smooth Muscle
Muscle
Myosin-Light-Chain Kinase
Phosphoserine
Carbachol
Phosphothreonine
Smooth Muscle Myosins
Light
Phosphorylation
Protein Kinase C
Phosphopeptides
Peptides
Peptide Mapping
Myosin Heavy Chains
Phosphates
Tissue
Muscles

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Colburn, J. C., Michnoff, C. H., Hsu, L. C., Slaughter, C. A., Kamm, K. E., & Stull, J. T. (1988). Sites phosphorylated in myosin light chain in contracting smooth muscle. Journal of Biological Chemistry, 263(35), 19166-19173.

Sites phosphorylated in myosin light chain in contracting smooth muscle. / Colburn, J. C.; Michnoff, C. H.; Hsu, L. C.; Slaughter, Clive A.; Kamm, K. E.; Stull, J. T.

In: Journal of Biological Chemistry, Vol. 263, No. 35, 01.12.1988, p. 19166-19173.

Research output: Contribution to journalArticle

Colburn, JC, Michnoff, CH, Hsu, LC, Slaughter, CA, Kamm, KE & Stull, JT 1988, 'Sites phosphorylated in myosin light chain in contracting smooth muscle', Journal of Biological Chemistry, vol. 263, no. 35, pp. 19166-19173.
Colburn JC, Michnoff CH, Hsu LC, Slaughter CA, Kamm KE, Stull JT. Sites phosphorylated in myosin light chain in contracting smooth muscle. Journal of Biological Chemistry. 1988 Dec 1;263(35):19166-19173.
Colburn, J. C. ; Michnoff, C. H. ; Hsu, L. C. ; Slaughter, Clive A. ; Kamm, K. E. ; Stull, J. T. / Sites phosphorylated in myosin light chain in contracting smooth muscle. In: Journal of Biological Chemistry. 1988 ; Vol. 263, No. 35. pp. 19166-19173.
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abstract = "Purified smooth muscle myosin light chain can be phosphorylated at multiple sites by myosin light chain kinase and protein kinase C. We have determined the sites phosphorylated on myosin light chain in intact bovine tracheal smooth muscle. Stimulation with 10 μM carbachol resulted in 66 ± 5{\%} monophosphorylated and 11 ± 2{\%} diphosphorylated myosin light chain after 1 min, and 47 ± 4{\%} monophosphorylated and 5 ± 2{\%} diphosphorylated myosin light chain after 30 min. Myosin heavy chain contained 0.06 ± 0.01 mol of phosphate/mol of protein which did not change with carbachol. At both 1 and 30 min the monophosphorylated myosin light chain contained only phosphoserine whereas the diphosphorylated myosin light chain contained both phosphoserine and phosphothreonine. Two-dimensional peptide mapping of tryptic digests of monophosphorylated and diphosphorylated myosin light chain obtained from carbachol-stimulated tissue was similar to the peptide maps of purified light chain monophosphorylated and diphosphorylated, respectively, by myosin light chain kinase; these maps were distinct from the map obtained with tracheal light chain phosphorylated by protein kinase C. Phosphorylation of tracheal smooth muscle myosin light chain by myosin light chain kinase yields the tryptic phosphopeptide ATSNVFAMFDQSQIQEFK with S the phosphoserine in the monophosphorylated myosin light chain and TS the phosphothreonine and phosphoserine in the diphosphorylated myosin light chain. Thus, stimulation of tracheal smooth muscle with a high concentration of carbachol results in formation of both monophosphorylated and diphosphorylated myosin light chain although the amount of diphosphorylated light chain is substantially less than monophosphorylated light chain. In the intact muscle, myosin light chain is phosphorylated at sites corresponding to myosin light chain kinase phosphorylation.",
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AU - Kamm, K. E.

AU - Stull, J. T.

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N2 - Purified smooth muscle myosin light chain can be phosphorylated at multiple sites by myosin light chain kinase and protein kinase C. We have determined the sites phosphorylated on myosin light chain in intact bovine tracheal smooth muscle. Stimulation with 10 μM carbachol resulted in 66 ± 5% monophosphorylated and 11 ± 2% diphosphorylated myosin light chain after 1 min, and 47 ± 4% monophosphorylated and 5 ± 2% diphosphorylated myosin light chain after 30 min. Myosin heavy chain contained 0.06 ± 0.01 mol of phosphate/mol of protein which did not change with carbachol. At both 1 and 30 min the monophosphorylated myosin light chain contained only phosphoserine whereas the diphosphorylated myosin light chain contained both phosphoserine and phosphothreonine. Two-dimensional peptide mapping of tryptic digests of monophosphorylated and diphosphorylated myosin light chain obtained from carbachol-stimulated tissue was similar to the peptide maps of purified light chain monophosphorylated and diphosphorylated, respectively, by myosin light chain kinase; these maps were distinct from the map obtained with tracheal light chain phosphorylated by protein kinase C. Phosphorylation of tracheal smooth muscle myosin light chain by myosin light chain kinase yields the tryptic phosphopeptide ATSNVFAMFDQSQIQEFK with S the phosphoserine in the monophosphorylated myosin light chain and TS the phosphothreonine and phosphoserine in the diphosphorylated myosin light chain. Thus, stimulation of tracheal smooth muscle with a high concentration of carbachol results in formation of both monophosphorylated and diphosphorylated myosin light chain although the amount of diphosphorylated light chain is substantially less than monophosphorylated light chain. In the intact muscle, myosin light chain is phosphorylated at sites corresponding to myosin light chain kinase phosphorylation.

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