Complete amino acid sequence of canine cardiac calsequestrin deduced by cDNA cloning

B. T. Scott, H. K.B. Simmerman, J. H. Collins, B. Nadal-Ginard, L. R. Jones

Research output: Contribution to journalArticle

165 Scopus citations

Abstract

cDNA cloning was used to deduce the complete amino acid sequence of canine cardiac calsequestrin, the principal Ca2+-binding protein of cardiac junctional sarcoplasmic reticulum. Cardiac calsequestrin contains 391 amino acid residues plus a 19-residue amino-terminal signal sequence. The molecular weight of the mature protein, excluding carbohydrate, is 45,269. Cardiac calsequestrin is highly acidic, and a striking feature is the enrichment of acidic residues (60%) within the 63 carboxyl-terminal residues. No part of the sequence contains EF and Ca2+-binding structures. The photoaffinity probe 3-(trifluoromethyl)-3-(m-[125I]iodophenyl)diazirine was used to localize the Ca2+-regulated hydrophobic site to amino acid residues 192-223. The cardiac and skeletal muscle isoforms of calsequestrin (Fliegel, L., Ohnishi, M., Carpenter, M.R., Khanna, V.K., Reithmeier, R.A.F., and MacLennan, D.H. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 1167-1171), although the products of different genes, are 65% identical, are acidic, and share one glcosylation site. However, cardiac calsequestrin has several unique features. First, it has a 31-amino acid extension at its carboxyl terminus (residues 361-391), which contains 71% acidic residues and a second glycosylation site. Second, its mRNA contains a second open reading frame with the capacity to code for a 111-amino acid protein. Third, contrary to the restricted expresion of the fast skeletal isoform, cardiac calsequestrin mRNA is present in both cardiac and slow skeletal muscle, but not in fast skeletal muscle. We conclude that the deduced amino acid sequence of cardiac calsequestrin is consistent with its ability to bind large amounts of Ca2+ (40 mol of Ca2+/mol of calsequestrin). The protein probably binds Ca2+ by acting as a charged surface rather than by presenting multiple discrete Ca2+-binding sites.

Original languageEnglish (US)
Pages (from-to)8958-8964
Number of pages7
JournalJournal of Biological Chemistry
Volume263
Issue number18
StatePublished - Jan 1 1988

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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