Purification and properties of the latent F1-APTase of Micrococcus lysodeikticus.
The latent coupling factor (F1)-ATPase of Micrococcus lysodeikticus has been purified to homogeneity as determined by a number of criteria including, nondenaturing polyacrylamide gel electrophoresis, crossed immunoelectrophoresis and analytical ultracentrifugation. By inclusion of 1 mM phenylmethyl sulfonyl fluoride, a serine protease inhibitor, in the shock-wash step of release of F1 from the membranes, the spontaneous activation of both crude and purified ATPase by endogenous membrane protease(s) can be prevented, thereby yielding a highly latent ATPase preparation. Equilibrium ultracentrifugation of the latent ATPase gave a molecular weight of 400 000. The ATPase contained five different subunits alpha, beta, gamma, delta, and espsilon and their molecular weights determined by SDS-polyacrylamide gel electrophoresis were 60 000, 54 000, 37 000, 27 000 and 9000, respectively. The subunit composition was determined with 14C-labelled, F1-ATPase prepared from cells grown on medium containing [U-14C]-labelled algal protein hydrolysate. Within the limitations of this method the results tentatively suggest a subunit composition of 3 : 3 : 1 : 1 : 3.
D000251:Adenosine Triphosphatases / Q000302:isolation & purification* / Q000378:metabolism; D000596:Amino Acids / Q000032:analysis; D007122:Immunoelectrophoresis; D046911:Macromolecular Substances; D008837:Micrococcus / Q000201:enzymology*; D008970:Molecular Weight; D010086:Oxidative Phosphorylation Coupling Factors / Q000302:isolation & purification* / Q000378:metabolism
D000596:Amino Acids; D046911:Macromolecular Substances; D010086:Oxidative Phosphorylation Coupling Factors; D000251:Adenosine Triphosphatases