A highly stable adenosine triphosphatase from a thermophillie bacterium. Purification, properties, and reconstitution.
1. A highly stable ATPase (TF1) was purified to a monodispersed state from the membranes of a thermophilic bacterium PS3. Its molecular weight was 380,000, and it was composed of five subunits alpha, beta, gamma, sigma', and sigma with molecular weights of 56,000, 53,000, 32,000, 15,500, and 11,000, respectively. 2. TF1 was stable against dissociating agents such as 5.5 M urea and 4.0 M LiCl, organic solvents, such as 60% acetone, heavy metals, and detergents. Low concentrations of all these agents stimulated its activity at 60 degrees. 3. TF1 was not cold-labile and showed a maximal activity at 70 degrees. Its CD spectrum revealed that its conformation changed between 81 and 96 degrees, and that its contents of alpha helices and beta structures were 27.3 and 12.8%, respectively, at 75 degrees. 4. TF1 was completely dissociated by treatment with dodecyl sulfate at 60 degrees and then with 7.1 M urea. The dissociated TF1 was reconstituted by treatment with Dowex 1-X2, and then dialysis. 5. [3H]Acetyl-TF1 bound to TF1-depleted membranes. TF1 only catalyzed 32Pi-ATP exchange and showed sensitivity to inhibitors of energy transfer when bound to the membranes. 6. A hydrophobic membrance component (TFo) was isolated which rendered TF1 sensitive to inhibitors of energy transfer. It was composed of three subunits (with molecular weights of 19,000, 13,500, and 5,400) and P-lipids.
D000251:Adenosine Triphosphatases / Q000302:isolation & purification / Q000378:metabolism*; D000596:Amino Acids / Q000032:analysis; D001419:Bacteria / Q000201:enzymology*; D002413:Cations, Divalent; D002462:Cell Membrane / Q000201:enzymology; D002942:Circular Dichroism; D004355:Drug Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D011487:Protein Conformation; D013696:Temperature
D000596:Amino Acids; D002413:Cations, Divalent; D046911:Macromolecular Substances; D000251:Adenosine Triphosphatases