Isolation of two isoforms of a 21,000-dalton Ca2+-binding protein of bovine brain.
Using Ca2+-dependent hydrophobic interaction chromatography we have identified a novel bovine brain Ca2+-binding protein (CaBP) composed of 21 kDa and 23 kDa polypeptides. This calciprotein was further purified by heat-treatment in the presence of Ca2+ and ion-exchange chromatography. The isolated protein exhibits a number of properties in common with proteins belonging to the calmodulin family of CaBPs, including a Ca2+-dependent electrophoretic mobility shift on SDS-polyacrylamide gel electrophoresis, retention of the ability to bind 45Ca2+ after electrophoresis and Western blotting, and a high content of acidic amino acids. We have recently isolated and characterized a 21 kDa CaBP from bovine brain and conclude that the 21 kDa and 21/23 kDa CaBPs are isoforms since they have very similar U.V. absorption spectra and amino acid compositions, and polyclonal antibodies raised in rabbits against the 21 kDa CaBP cross-react to an identical degree with the 21/23 kDa CaBP as determined by the competitive enzyme-linked immunosorbent assay (ELISA). Both proteins contain carbohydrate, but they differ in the degree of glycosylation. Tissue distribution studies indicate the presence of both 21 kDa and 23 kDa Ca2+-binding polypeptides in bovine trachea, aorta, kidney, skeletal muscle and cardiac muscle, and chicken gizzard smooth muscle.
Department of Medical Biochemistry, University of Calgary, Alberta, Canada.
J R
JR
McDonald
U
U
Gröschel-Stewart
M P
MP
Walsh
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001921:Brain / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002132:Calcium Radioisotopes; D002135:Calcium-Binding Proteins / Q000302:isolation & purification* / Q000378:metabolism; D002147:Calmodulin / Q000302:isolation & purification / Q000378:metabolism; D002417:Cattle; D002852:Chromatography, Ion Exchange; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D008970:Molecular Weight
D000596:Amino Acids; D002132:Calcium Radioisotopes; D002135:Calcium-Binding Proteins; D002147:Calmodulin; D002118:Calcium