Purification and distribution of a novel macrophage-specific calmodulin-binding glycoprotein

S. J. Orlow, David L. Rosenstreich, S. Pifko-Hirst, O. M. Rosen

Research output: Contribution to journalArticle

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Abstract

The murine macrophage-like cell line J774.16 and peritoneal exudate cells elicited with thioglycollate or starch contain a major calmodulin (CaM)-binding protein (CaMBP) which is absent in trifluoroperazine-resistant variants of J774, resident peritoneal macrophages and peritoneal macrophages elicited with concanavalin A, lipopolysaccharide, proteose peptone, or Bacillus Calmette-Guerin. Resident murine peritoneal cells maintained in tissue culture for 3 days begin to accumulate this protein, as do human peripheral blood monocytes after 7 days of culture. A specific competitive displacement radioimmunoassay was developed with the use of a rabbit antiserum raised to the partially purified CaM-binding protein and [125I]CaM covalently cross-linked to the principal CaM-binding protein in the preparation. The radioimmunoassay confirmed the unique cellular distribution of this protein, suggesting that it may be a marker for certain stages of macrophage differentiation. Monoclonal antibodies were prepared, and one of these was used to further purify the protein by immunoaffinity chromatography. A protein of M(r) 50,000 to 60,000 was isolated. The protein could be selectively adsorbed to wheat germ agglutinin agarose and subsequently eluted with N-acetyl glucosamine. This property, plus the sensitivity of the protein to endoglycosidase F, led to the conclusion that it is a glycoprotein. The cellular distribution, subcellular localization, and evidence for glycosylation suggest that this protein may be a macrophage-specific receptor with a high affinity for Ca2+ -CaM.

Original languageEnglish (US)
Pages (from-to)449-456
Number of pages8
JournalJournal of Immunology
Volume134
Issue number1
StatePublished - 1985

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Calmodulin
Glycoproteins
Macrophages
Calmodulin-Binding Proteins
Proteins
Peritoneal Macrophages
Radioimmunoassay
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase
Thioglycolates
Trifluoperazine
Wheat Germ Agglutinins
Glucosamine
Exudates and Transudates
Mycobacterium bovis
Concanavalin A
Glycosylation
Starch
Sepharose
Lipopolysaccharides
Chromatography

ASJC Scopus subject areas

  • Immunology

Cite this

Purification and distribution of a novel macrophage-specific calmodulin-binding glycoprotein. / Orlow, S. J.; Rosenstreich, David L.; Pifko-Hirst, S.; Rosen, O. M.

In: Journal of Immunology, Vol. 134, No. 1, 1985, p. 449-456.

Research output: Contribution to journalArticle

Orlow, S. J. ; Rosenstreich, David L. ; Pifko-Hirst, S. ; Rosen, O. M. / Purification and distribution of a novel macrophage-specific calmodulin-binding glycoprotein. In: Journal of Immunology. 1985 ; Vol. 134, No. 1. pp. 449-456.
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