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Purification, characterization, and localization of 70 kDa calcium‐sensitive protein (calelectrin) from mammary glands
Author(s) -
Lozano Joseph J.,
Haindl Alfred H.,
Rocha Victor
Publication year - 1989
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.1041410213
Subject(s) - proteolysis , biology , microbiology and biotechnology , calcium , antigenicity , mammary gland , biochemistry , immunohistochemistry , antigen , chemistry , enzyme , immunology , genetics , organic chemistry , cancer , breast cancer
Mammary glands contain a group of calcium‐sensitive proteins that bind to membranes in a calcium‐dependent manner. Using the calcium‐dependent binding to hydrophobic surfaces in combination with conventional techniques, we have purified the 70kDa mammary calcium‐binding protein (70 kDa M‐CBP) to homogeneity. Antisera prepared to the 70 kDa M‐CBP or to bovine liver 67 kDa calelectrin reacted in immunoblot analysis with the 70 kDa M‐CBP antigen and with several additional mammary CBP species in crude tissue homogenates. Limited proteolysis of the 70 kDa M‐CBP produced smaller immunoreactive species; extensive proteolysis resulted in more complete degradation of the protein. Identical data were obtained with digestion of 67 kDa calelectrin. The pi for the 70 kDa M‐CBP was determined to be approximately 5.8; the same value reported for 67 kDa calelectrin. Phosphorylation of 70 kDa M‐CBP was not detected in epithelial cell culture metabolic labeling. Immunohistochemical localization showed the protein to be located in ductal epithelia of virgin mouse mammary glands with a pattern of increased staining of the basal portions of the cells. Some stromal cells were also reactive. Apparently, the 70 kDa M‐CBP and 67 kDa calelectrin are the same protein. Furthermore, like the 32.5 calelectrin (endonexin) and calpactin 1/p36/lipocortin II, the 70 kDa protein appears to be a ductal epithelial cell associated protein in the mammary gland.