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Calmodulin binding domains: characterization of a phosphorylation and calmodulin binding site from myosin light chain kinase

A protein kinase phosphorylation site in chicken gizzard myosin light chain kinase (MLCK) has been identified, and a synthetic peptide analogue of this site has been shown to be a high-affinity calmodulin binding peptide as well as a substrate for cyclic AMP dependent protein kinase. Phosphorylation...

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Published in:Biochemistry (Easton) 1986-03, Vol.25 (6), p.1458-1464
Main Authors: Lukas, Thomas J, Burgess, Wilson H, Prendergast, Franklyn G, Lau, Wai, Watterson, D. Martin
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container_issue 6
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container_title Biochemistry (Easton)
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creator Lukas, Thomas J
Burgess, Wilson H
Prendergast, Franklyn G
Lau, Wai
Watterson, D. Martin
description A protein kinase phosphorylation site in chicken gizzard myosin light chain kinase (MLCK) has been identified, and a synthetic peptide analogue of this site has been shown to be a high-affinity calmodulin binding peptide as well as a substrate for cyclic AMP dependent protein kinase. Phosphorylation of the site in MLCK is diminished when reactions are done in the presence of calmodulin. A fragment of MLCK containing the phosphorylation site was shown to have the amino acid sequence Ala-Arg-Arg-Lys-Trp-Gln-Lys-Thr-Gly-His-Ala-Val-Arg-Ala-Ile-Gly-Arg-Leu- Ser-Ser. The interaction of calmodulin with a synthetic peptide based on this sequence was characterized by using circular dichroism and fluorescence spectroscopies and inhibition of calmodulin activation of MLCK. The peptide-calmodulin complex had an estimated dissociation constant in the range of 1 nM, underwent spectroscopic changes in the presence of calmodulin consistent with the induction of an alpha-helical structure, and interacted with calmodulin with an apparent 1:1 stoichiometry. Studies with other synthetic peptide analogues indicated that the phosphorylation of the serine residues diminished the ability of the peptide to interact with calmodulin even though the serines are not required for calmodulin binding. On the basis of the primary and secondary structural characteristics of these peptide analogues, a potential calmodulin binding region in another calmodulin binding protein, the gamma subunit of rabbit skeletal muscle phosphorylase kinase, was identified.
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Martin</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Calmodulin binding domains: characterization of a phosphorylation and calmodulin binding site from myosin light chain kinase</atitle><jtitle>Biochemistry (Easton)</jtitle><addtitle>Biochemistry</addtitle><date>1986-03-01</date><risdate>1986</risdate><volume>25</volume><issue>6</issue><spage>1458</spage><epage>1464</epage><pages>1458-1464</pages><issn>0006-2960</issn><eissn>1520-4995</eissn><abstract>A protein kinase phosphorylation site in chicken gizzard myosin light chain kinase (MLCK) has been identified, and a synthetic peptide analogue of this site has been shown to be a high-affinity calmodulin binding peptide as well as a substrate for cyclic AMP dependent protein kinase. Phosphorylation of the site in MLCK is diminished when reactions are done in the presence of calmodulin. A fragment of MLCK containing the phosphorylation site was shown to have the amino acid sequence Ala-Arg-Arg-Lys-Trp-Gln-Lys-Thr-Gly-His-Ala-Val-Arg-Ala-Ile-Gly-Arg-Leu- Ser-Ser. The interaction of calmodulin with a synthetic peptide based on this sequence was characterized by using circular dichroism and fluorescence spectroscopies and inhibition of calmodulin activation of MLCK. The peptide-calmodulin complex had an estimated dissociation constant in the range of 1 nM, underwent spectroscopic changes in the presence of calmodulin consistent with the induction of an alpha-helical structure, and interacted with calmodulin with an apparent 1:1 stoichiometry. Studies with other synthetic peptide analogues indicated that the phosphorylation of the serine residues diminished the ability of the peptide to interact with calmodulin even though the serines are not required for calmodulin binding. 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identifier ISSN: 0006-2960
ispartof Biochemistry (Easton), 1986-03, Vol.25 (6), p.1458-1464
issn 0006-2960
1520-4995
language eng
recordid cdi_proquest_miscellaneous_76789043
source ACS CRKN Legacy Archives
subjects Amino Acids - analysis
Analytical, structural and metabolic biochemistry
Animals
Binding Sites
Biological and medical sciences
calmodulin
Calmodulin - metabolism
Chickens
Enzymes and enzyme inhibitors
Fundamental and applied biological sciences. Psychology
gizzard
Gizzard, Avian - enzymology
Kinetics
Macromolecular Substances
Molecular Weight
Muscle, Smooth - enzymology
Myosin-Light-Chain Kinase
Peptide Fragments - analysis
Phosphopeptides - analysis
Phosphorylation
Protein Binding
Protein Conformation
Protein Kinases - metabolism
Transferases
title Calmodulin binding domains: characterization of a phosphorylation and calmodulin binding site from myosin light chain kinase
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