Sequence of subunit c of the Na⁺‐translocating F₁F_O ATPase of Acetobacterium woodii: proposal for determinants of Na⁺ specificity as revealed by sequence comparisons

Abstract

A 3.2 kb EcoRl fragment carrying genes for Na + -FjFo ATPase was cloned from chromosomal DNA of Acetobacterium woodii. DNA sequence analysis revealed the presence of an open reading frame which was identified by data base searches and comparison with the experimentally derived Nterminal amino acid sequence to code for subunit c of Na + -F 1 F 0 ATPase. A comparison of the primary sequences of the two well established Na + -translocating F^o ATPases from Acetobacterium woodii and Propionigenium modestum with H + -translocating enzymes indicates the length of the C-terminus as well as specific residues located in the cytoplasmic membrane to be important for Na + transport.

Experimental Verification of a Sequence-Based Prediction: F 1 F 0 -Type ATPase of Vibrio cholerae Transports Protons, Not Na + Ions

“…Although vibrial AtpE subunits had longer C-terminal fragments than did the H ϩ ATPases from E. coli and B. subtilis and the Na ϩ ATPases from A. woodii and P. modestum, they clearly lacked the predicted Na ϩ -binding motif (Table 1). This apparent contradiction of the two previously established criteria of ATPase cation specificity (27) prompted us to investigate the nature of the coupling ion in V. cholerae F 1 F 0 ATPase in more detail and find out whether V. cholerae uses the sodium motive or proton motive force in oxidative phosphorylation.…”

“…The acidic (Asp or Glu) residue in this position is conserved among c subunits of both H ϩ -dependent and Na ϩ -dependent F 1 F 0 ATPases from various bacteria, as well as among the equivalent K subunits of the archaeal-and vacuolar-type (A/V-type) ATPases (reviewed in reference 1) ( Table 1). In Na ϩ -conducting c and K subunits, however, the Glu residue is followed by a hydroxylcontaining (Ser or Thr) residue, which apparently provides additional liganding groups, which are essential for binding alkali cations (20,27). The presence of conserved Pro and Gln residues on the adjacent transmembrane segment and the overall membrane topology of the c subunit have also been implicated in the determination of the cation selectivity of the enzyme (19,20,27).…”

“…In Na ϩ -conducting c and K subunits, however, the Glu residue is followed by a hydroxylcontaining (Ser or Thr) residue, which apparently provides additional liganding groups, which are essential for binding alkali cations (20,27). The presence of conserved Pro and Gln residues on the adjacent transmembrane segment and the overall membrane topology of the c subunit have also been implicated in the determination of the cation selectivity of the enzyme (19,20,27). Combining the available data, Rahlfs and Müller (27) proposed that there are two determinants of Na ϩ specificity for the F 1 F 0 ATPase of A. woodii: (i) an enlargement of the C terminus of subunit c and (ii) the presence of the Na ϩ -binding motif of P 25 , Q 29 , E 62 , and T 63 (Table 1).…”

“…The presence of conserved Pro and Gln residues on the adjacent transmembrane segment and the overall membrane topology of the c subunit have also been implicated in the determination of the cation selectivity of the enzyme (19,20,27). Combining the available data, Rahlfs and Müller (27) proposed that there are two determinants of Na ϩ specificity for the F 1 F 0 ATPase of A. woodii: (i) an enlargement of the C terminus of subunit c and (ii) the presence of the Na ϩ -binding motif of P 25 , Q 29 , E 62 , and T 63 (Table 1). An inspection of the AtpE sequences from V. alginolyticus (23) and V. cholerae (16) showed that they share 92% identity and are very similar to the H ϩ -conducting c subunits of the F 1 F 0 enzymes from E. coli, Bacillus subtilis, Enterococcus hirae, and mitochondria of Saccharomyces cerevisiae and humans (Table 1).…”

Biochemical and Molecular Characterization of a Na + -Translocating F 1 F o -ATPase from the Thermoalkaliphilic Bacterium Clostridium paradoxum

“…8) showed stronger overall homology with Na ϩ -translocating F 1 F o -ATP synthases from A. woodii (83% identity), P. modestum (67% identity), and I. tartaricus (70% identity) than with the proton-coupled E. coli counterpart (27% identity) or those from other clostridia (42 to 50% identity). Furthermore, the Na ϩ -binding motif consisting of Q 28 , E 61 , and S 62 (21,37,39,45) was present in the c subunit of the C. paradoxum ATP synthase.…”