Na+-translocating membrane pyrophosphatases are widespread in the microbial world and evolutionarily precede H+-translocating pyrophosphatases

J Biol Chem. 2011 Jun 17;286(24):21633-42. doi: 10.1074/jbc.M111.244483. Epub 2011 Apr 28.


Membrane pyrophosphatases (PPases), divided into K(+)-dependent and K(+)-independent subfamilies, were believed to pump H(+) across cell membranes until a recent demonstration that some K(+)-dependent PPases function as Na(+) pumps. Here, we have expressed seven evolutionarily important putative PPases in Escherichia coli and estimated their hydrolytic, Na(+) transport, and H(+) transport activities as well as their K(+) and Na(+) requirements in inner membrane vesicles. Four of these enzymes (from Anaerostipes caccae, Chlorobium limicola, Clostridium tetani, and Desulfuromonas acetoxidans) were identified as K(+)-dependent Na(+) transporters. Phylogenetic analysis led to the identification of a monophyletic clade comprising characterized and predicted Na(+)-transporting PPases (Na(+)-PPases) within the K(+)-dependent subfamily. H(+)-transporting PPases (H(+)-PPases) are more heterogeneous and form at least three independent clades in both subfamilies. These results suggest that rather than being a curious rarity, Na(+)-PPases predominantly constitute the K(+)-dependent subfamily. Furthermore, Na(+)-PPases possibly preceded H(+)-PPases in evolution, and transition from Na(+) to H(+) transport may have occurred in several independent enzyme lineages. Site-directed mutagenesis studies facilitated the identification of a specific Glu residue that appears to be central in the transport mechanism. This residue is located in the cytoplasm-membrane interface of transmembrane helix 6 in Na(+)-PPases but shifted to within the membrane or helix 5 in H(+)-PPases. These results contribute to the prediction of the transport specificity and K(+) dependence for a particular membrane PPase sequence based on its position in the phylogenetic tree, identity of residues in the K(+) dependence signature, and position of the membrane-located Glu residue.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biological Transport
  • Cations
  • Cytoplasm / metabolism
  • Dose-Response Relationship, Drug
  • Escherichia coli / enzymology
  • Evolution, Molecular
  • Hydrolysis
  • Inorganic Pyrophosphatase / metabolism*
  • Kinetics
  • Ligands
  • Potassium / chemistry
  • Proton Pumps
  • Protons
  • Recombinant Proteins / chemistry
  • Sodium / chemistry
  • Sodium / metabolism


  • Cations
  • Ligands
  • Proton Pumps
  • Protons
  • Recombinant Proteins
  • Sodium
  • Inorganic Pyrophosphatase
  • Potassium