The Na ؉ -F 1 F 0 -ATPase operon of Acetobacterium woodii was recently shown to contain, among eleven atp genes, those genes that encode subunit a and b, a gene encoding a 16-kDa proteolipid (subunit c 1 ), and two genes encoding 8-kDa proteolipids (subunits c 2 and c 3 ). Because subunits a, b, and c 1 were not found in previous enzyme preparations, we re-determined the subunit composition of the enzyme. The genes were overproduced, and specific antibodies were raised. Western blots revealed that subunits a, b, and c 1 are produced and localized in the cytoplasmic membrane. Membrane protein complexes were solubilized by dodecylmaltoside and separated by blue native-polyacrylamide gel electrophoresis, and the ATPase subunits were resolved by SDS-polyacrylamide gel electrophoresis. N-terminal sequence analyses revealed the presence of subunits a, c 2 , c 3 , b, ␦, ␣, ␥, , and ⑀. Biochemical and immunological analyses revealed that subunits c 1 , c 2 , and c 3 are all part of the c-oligomer, the first of a F 1 F 0 -ATPase that contains 8-and 16-kDa proteolipids.