Identification of novel calcium binding proteins of heart and brain 100,000 x g supernatant

Abstract

The chelex competitive calcium binding assay has been used to assay the calcium binding activity of the 100,000 x g supernatant of bovine heart and brain. Chromatography of brain 100,000 x g supernatant on diethylaminoethyl (DEAE) cellulose reveals the presence of two peaks of calcium binding activity, peak I eluting at about 0.05 M NaCl and peak II at about 0.18 M NaCl. Chromatography of peak I on Sephadex G-150 resolves a major and a minor peak of calcium binding activity, at M r 40,000 and M r 150,000. Chromatography of peak II (0.18 M NaCl) on Sepharose 6B produces two peaks of calcium binding activity, a broad peak of calcium binding activity composed of two molecular weight species of M r 230,000 and M r 420,000, and a sharp peak of calcium binding activity with M r 75,000. Chromatography of the 100,000 x g supernatant of bovine heart on DEAE Cellulose reveals two peaks of calcium binding activity. Chromatography of the lower ionic strength peak on Sephadex G-150 resolved major and minor peaks of calcium binding activity at M r 65,000 and 150,000, respectively. The results of this study suggest the presence of several calcium binding proteins, other than calmodulin, in these tissues.