Purification of a 90-kDa protein (band VII) from cardiac sarcoplasmic reticulum

Identification as calnexin and localization of casein kinase II phosphorylation sites

Steven E. Cala, Corinne Ulbright, Jeffry S. Kelley, Larry Jones

Research output: Contribution to journalArticle

38 Citations (Scopus)

Abstract

Purification and sequencing of proteins from cardiac sarcoplasmic reticulum (SR) vesicles have provided a framework for the study of SR function. Included among the SR proteins so far investigated are a collection of intralumenal proteins that stain blue with the protein dye Stains-All in sodium dodecyl sulfate-polyacrylamide gels. A single prominent blue staining SR protein of apparent Mr = 90,000 (Band VII), however, has remained uncharacterized. In the work described here, purification of Band VII from dog cardiac SR vesicles, along with amino acid sequencing and cDNA cloning, identified this protein as calnexin, a homologue of calreticulin recently described in dog pancreatic microsomes. Using ryanodine-mediated calcium oxalate loading of SR vesicles followed by density gradient centrifugation, we have shown that calnexin is a bona fide SR protein and an integral constituent of both junctional and free SR vesicles. Calnexin was found to be a substrate for casein kinase II and was phosphorylated at two distinct sites localized to the carboxyl- and amino-terminal ends of the molecule. Enrichment of calnexin in cardiac SR vesicles indicates a role for calnexin involving the specialized function of the SR membrane.

Original languageEnglish
Pages (from-to)2969-2975
Number of pages7
JournalJournal of Biological Chemistry
Volume268
Issue number4
StatePublished - Feb 5 1993

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Calnexin
Casein Kinase II
Phosphorylation
Sarcoplasmic Reticulum
Purification
Proteins
Coloring Agents
Protein Sequence Analysis
Calreticulin
Ryanodine
Calcium Oxalate
Centrifugation
Cloning
Dogs
Sodium Dodecyl Sulfate
Density Gradient Centrifugation
Complementary DNA
Microsomes
Membranes
Amino Acids

ASJC Scopus subject areas

  • Biochemistry

Cite this

Purification of a 90-kDa protein (band VII) from cardiac sarcoplasmic reticulum : Identification as calnexin and localization of casein kinase II phosphorylation sites. / Cala, Steven E.; Ulbright, Corinne; Kelley, Jeffry S.; Jones, Larry.

In: Journal of Biological Chemistry, Vol. 268, No. 4, 05.02.1993, p. 2969-2975.

Research output: Contribution to journalArticle

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