Purification of a novel calmodulin binding protein from bovine cerebral cortex membranes

Terrence J. Andreasen, Charles W Luetje, Warren Heideman, Daniel R. Storm

Research output: Contribution to journalArticle

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Abstract

A new calmodulin (CaM) binding protein, designated P-57, has been purified to apparent homogeneity from bovine cerebral cortex membranes. In contrast to other calmodulin binding proteins, P-57 has higher affinity for calmodulin in the absence of bound Ca2+ than in its presence. The protein was purified by DEAE-Sephacel chromatography and two CaM-Sepharose affinity column steps. The first CaM-Sepharose column was run in the presence of Ca2+; the second was run in the presence of chelator in excess of Ca2+. P-57 was adsorbed by CaM-Sepharose only in the absence of bound Ca2+ and was eluted from the second column by buffers containing Ca2+. Sodium dodecyl sulfate (SDS)-polyacrylamide gels of the purified protein showed only one band at Mr 57 000. The major form of the protein on Bio-Gel A-1.5m and native polyacrylamide gradient gel electrophoresis ran with an apparent Stokes radius of 41 Å. Photoaffinity labeling of P-57 with azido[125I]calmodulin yielded one cross-linked product on SDS gels with an Mr of 70 000. This interaction occurred only when excess ethylene glycol bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid was present and was inhibited by the presence of Ca2+ in excess of chelator. It appears that P-57 has novel binding properties for calmodulin distinct from all other calmodulin binding proteins described thus far.

Original languageEnglish
Pages (from-to)4615-4618
Number of pages4
JournalBiochemistry
Volume22
Issue number20
StatePublished - Dec 1 1983
Externally publishedYes

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Calmodulin-Binding Proteins
Calmodulin
Cerebral Cortex
Purification
Membranes
Sepharose
Gels
Chelating Agents
Sodium Dodecyl Sulfate
Native Polyacrylamide Gel Electrophoresis
Proteins
Ethylene Glycol
Chromatography
Electrophoresis
Ether
Labeling
Buffers
Acids

ASJC Scopus subject areas

  • Biochemistry

Cite this

Andreasen, T. J., Luetje, C. W., Heideman, W., & Storm, D. R. (1983). Purification of a novel calmodulin binding protein from bovine cerebral cortex membranes. Biochemistry, 22(20), 4615-4618.

Purification of a novel calmodulin binding protein from bovine cerebral cortex membranes. / Andreasen, Terrence J.; Luetje, Charles W; Heideman, Warren; Storm, Daniel R.

In: Biochemistry, Vol. 22, No. 20, 01.12.1983, p. 4615-4618.

Research output: Contribution to journalArticle

Andreasen, TJ, Luetje, CW, Heideman, W & Storm, DR 1983, 'Purification of a novel calmodulin binding protein from bovine cerebral cortex membranes', Biochemistry, vol. 22, no. 20, pp. 4615-4618.
Andreasen TJ, Luetje CW, Heideman W, Storm DR. Purification of a novel calmodulin binding protein from bovine cerebral cortex membranes. Biochemistry. 1983 Dec 1;22(20):4615-4618.
Andreasen, Terrence J. ; Luetje, Charles W ; Heideman, Warren ; Storm, Daniel R. / Purification of a novel calmodulin binding protein from bovine cerebral cortex membranes. In: Biochemistry. 1983 ; Vol. 22, No. 20. pp. 4615-4618.
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