Proton Translocating ATPase of a Thermophilic Bacterium

Abstract

1. A stable membrane-bound ATPase [EC 3.6. 1. 3] (TF₀.F₁) capable of proton translocation in reconstituted vesicles was purified from the thermophilic bacterium PS3 cultured in medium containing L-[U-₁₄C]amino acids. 2. TF₀.F₁ was composed of a catalytic moiety (TF₁) and a hydrophobic moiety (TF₀). TF₁ contained 3 polypeptide chains with molecular weights of 56,000, 3 of 53,000, 1 of 32,000, 1 of 15,500, and 1 of 11,000. TF₀ contained 1 chain of 19,000, 2 of 13,500, and 5 of 5,400 daltons. TF₁ was dissociated into subunits much less readily than F₁. 3. TF₁ consisted of 95A particles arrayed in hexagonal microcrystals. TF₀. F₁ consisted of a sphere (TF₁) and a stalk plus base (TF₀) which was buried in the membrane of the proton translocating vesicles. 4. Vesicles capable of energy transformation were formed when TF₁ came in contact with the surface of liposomes containing TF_o. On addition of phospholipids, the helix content of TF₀ increased 3-fold. The role of F_o in forming channels for protons is discussed. 5. The amino acid compositions of TF₀, TF₁, and TF₀.F₁ were compared. TF₀ was not hydrophobic, despite its interaction with phospholipids. The phospholipid composition and other properties of the proton translocating vesicles were examined. Vesicles reconstituted from a mixture of phosphatidylethanolamine, phosphatidylgly-cerol, and cardiolipin in the same ratio as in the membranes had the highest activity.