title: Purification and characterization of two forms of a low-affinity Ca2+-ATPase from erythrocyte membranes.
abstract: A low-affinity Ca2+-ATPase from erythrocyte membranes has been purified by agarose suspension electrophoresis and polyacrylamide gel electrophoresis in the absence of detergents. For maximal activity a calcium concentration above 10 mM is required. The activity is independent of magnesium. The Km value for ATP is about 60 microM. The enzyme appears in two forms (A and B) with similar amino acid composition. The specific activity of A is higher than that of B. Gel electrophoresis in SDS of A gives a pattern consisting of two bands. B gives the same pattern; the only difference between the patterns is the ratio of the amounts of protein in the bands. The apparent molecular weight of the proteins in the two SDS bands has been estimated at 23000 and 21000, respectively. The results obtained can be explained by assuming that the two proteins corresponding to the two bands obtained in SDS electrophoresis have a similar structure and can associate to complexes A and B. We have also shown that electrophoretic and chromatographic supporting media can induce aggregation of (membrane) proteins. Artificial complexes can thus be formed and cause misinterpretation of the data obtained. This may be the reason why some authors have speculated that Ca2+-ATPase is active only in complex with other proteins such as spectrin and actin.
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000596:Amino Acids / Q000032:analysis; D002118:Calcium / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000097:blood* / Q000302:isolation & purification; D004910:Erythrocyte Membrane / Q000201:enzymology*; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008970:Molecular Weight; D011487:Protein Conformation
chemical_list: D000596:Amino Acids; D000255:Adenosine Triphosphate; D000252:Calcium-Transporting ATPases; D008274:Magnesium; D002118:Calcium