Dual mode of energy coupling by the oxyanion-translocating ArsB protein

Abstract

The arsA and arsB genes of the ars operon of R-factor R773 confer arsenite resistance in Escherichia coli by coding for an anion-translocating ATPase. Arsenite resistance and the in vivo energetics of arsenite transport were compared in cells expressing the arsA and arsB genes and those expressing just the arsB gene. Cells expressing the arsB gene exhibited intermediate arsenite resistance compared with cells expressing both the arsA and arsB genes. Both types of cells exhibited energy-dependent arsenite exclusion. Exclusion of 73 AsO 2 ؊ from cells expressing only the arsB gene was coupled to electrochemical energy, while in cells expressing both genes, transport was coupled to chemical energy, most likely ATP. These results suggest that the Ars anion transport system can be either an obligatory ATP-coupled primary pump or a secondary carrier coupled to the proton motive force, depending on the subunit composition of the transport complex.Bacterial resistance to arsenical and antimonial compounds results from active extrusion of the toxic oxyanions from the resistant cells (15). Efflux of arsenite from oxyanion resistant Escherichia coli cells is catalyzed by a pump whose genes are encoded on the conjugative R factor R773 (5, 10). This aniontranslocating ATPase is composed of two types of polypeptides subunits, the ArsA and ArsB proteins. The 63-kDa ArsA protein is an anion-stimulated ATPase (11, 23) that functions as the catalytic component of the pump and remains attached to the inner membrane through interaction with the ArsB subunit (7, 30). The ArsB polypeptide is a 45-kDa inner membrane protein (26) that presumably forms the anion-conducting pathway.In vivo chemical energy was shown to be both necessary and sufficient for the pump, while electrochemical energy was neither (18,22). These studies were performed with an unc strain of E. coli defective in the H ϩ -translocating ATPase that catalyzes the equilibrium between ATP and the electrochemical proton gradient. This allows the establishment of intracellular conditions such that either chemical energy or electrochemical energy is available for transport (2). While the identity of the chemical used for energy coupling cannot be determined from the in vivo results, the energy requirement in vitro was similarly linked to chemical energy. In everted membrane vesicles, ATP was specifically required for uptake of arsenite, while electrochemical energy was neither necessary nor sufficient (6). Thus, the R773 ArsA-ArsB complex is clearly an obligatorily ATPcoupled pump.Similar arsenite resistance operons have been identified in plasmids of the gram-positive bacteria Staphylococcus aureus(pI258) (12) and Staphylococcus xylosus(pSX267) (24). The staphylococcal ars operons do not contain an arsA gene, yet the resistant cells still actively extrude arsenite (27,28), suggesting that the ArsB protein can transport arsenite in the absence of an arsA gene product. It is possible that the cells use a chromosomally encoded ATPase as a replacement for the ArsA protein or t...

The Saccharomyces cerevisiae ACR3 Gene Encodes a Putative Membrane Protein Involved in Arsenite Transport

“…The Acr3p probably extrudes unconjugated oxyanions of arsenite out of cells, thus reducing the intracellular concentration of toxic anions and providing resistance phenotype. This transport is probably coupled to the protonmotive force as it was suggested for the ArsB-dependent arsenite efflux which correlated to the membrane potential and not to the ATP content (3,23). We believe that As(V) is first reduced to As(III) by an arsenate reductase prior to the excretion by the Acr3p.…”

“…When the ArsA protein catalytic subunit is absent, the transport of oxyanions is carried out through the ArsB protein coupled to the protonmotive force (23). Prokaryotic cells extrude arsenicals only in the form of unconjugated arsenite (23,FIG. 5.…”

Arsenite transport by mammalian aquaglyceroporins AQP7 and AQP9

“…Arsenite and antimonite are commonly considered oxyanions. Indeed, transport of arsenite by the bacterial ArsB carrier is driven by the membrane potential, suggesting anionic substrates (33,34). However the trivalent arsenic and antimony acids have pKa values of 9.2 and 11.8, respectively, so that at neutral pH they would be primarily present in solution as neutral species, As(OH) 3 or Sb(OH) 3 .…”

Diversity of transport mechanisms: common structural principles

“…The ArsAB complex is quite stable, dissociating only in the presence of chaotropic agents. Second, when the in vivo energetic experiments were performed in cells expressing both the arsA and arsB genes, quite different results were obtained, which clearly demonstrate that the ArsAB complex is an obligatorily ATP-coupled primary transporter 22 . In an unc strain of E. coli that expressed both ArsA and ArsB, succinate no longer supports arsenite extrusion, and glucose-coupled transport is insensitive to uncouplers and respiratory chain inhibitors.…”

“…From a combination of in vivo and in vitro studies, which are outlined below, it is clear that arsenite transport catalysed by ArsB requires only the pmf and not ATP, suggesting that ArsB is a secondary transporter that extrudes the oxyanion out of cells 22,23 . In cells expressing arsB, transport requires the pmf and is inhibited by uncouplers 22 . For these studies, an unc strain of E. coli lacking the H ϩ -translocating ATPase that catalyses the equilibrium between ATP and the pmf was used.…”