Protein-protein interaction of detergent-solubilized Ca2(+)-ATPase during ATP hydrolysis analyzed by low-angle laser light scattering photometry coupled with high-performance gel chromatography.
Protein-protein interaction of detergent-solubilized Ca2(+)-ATPase was examined, employing low-angle laser light scattering photometry coupled with high-performance gel chromatography. When solubilized with octa(ethylene glycol) mono-n-dodecyl ether (C12E8) and chromatographed in the presence of 0.3 mg/ml C12E8, the Ca2(+)-ATPase emerged as a single peak with an intermediate molecular weight between the monomer and the dimer, showing a dissociation-association equilibrium of the two components. In the presence of 50 micrograms/ml phosphatidylcholine and 0.3 mg/ml C12E8 at 0 degrees C, the Ca2(+)-ATPase (0.8 mg) emerged as the two distinct components with molecular weights of 125,000 +/- 2100 (n = 3) and 211 300 +/- 7300 (n = 3), indicating that there was no rapid interconversion between the monomer and the dimer. Under the latter conditions, addition of ATP induced fusion of two components. The apparent molecular weight of the fused peak shifted from the monomer to the dimer as the amount of protein increased. Addition of ADP or adenosine 5'-(beta, gamma-methylene triphosphate), however, did not induce such fusion of the peaks. The ATP-induced fusion of the peaks was not observed either in 5 mM CaCl2, the conditions in which the rate of ATP hydrolysis was extremely slow. Thus, the solubilized Ca2(+)-ATPase underwent a rapid interconversion between the monomer and the dimer during ATP hydrolysis. These results suggest that the protein-protein interaction during ATP hydrolysis is an intrinsic nature of Ca2(+)-ATPase and that such interaction may be important for Ca2+ transport by Ca2(+)-ATPase in the sarcoplasmic reticulum membranes.
Kijima
Y
Y
First Department of Medicine, Osaka University School of Medicine, Japan.
Takagi
T
T
Shigekawa
M
M
Tada
M
M
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D000252:Calcium-Transporting ATPases / Q000302:isolation & purification / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D003902:Detergents; D006868:Hydrolysis; D007834:Lasers; D046911:Macromolecular Substances; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology; D012542:Scattering, Radiation; D012995:Solubility
D003902:Detergents; D046911:Macromolecular Substances; D000255:Adenosine Triphosphate; D000252:Calcium-Transporting ATPases