The Na⁺ ‐motive terminal oxidase activity in an alkalo‐ and halo‐tolerant Bacillus

Abstract

An alkalo-and halo-tolerant aerobic microorganism has been isolated which, according to microbiological analysis data and the ribosomal 5s RNA sequence, is a Bacillus similar, but not identical, to B. lichenijormis and B. subtilis. The microorganism, called Bacillus FTU, proved to be resistant to the protonophorous uncoupler carbonylcyanide m-chlorophenylhydrazone (CCCP). The fast growth of Bacillus FTU in the presence of CCCP was shown to require a high Na' concentration in the medium. A procedure was developed to exhaust endogenous respiratory substrates in Bacillus FTU cells so that fast oxygen consumption by the cells was observed only when an exogenous respiratory substrate was added. The exhausted cells were found to oxidize ascorbate in the presence of N,N,N,N'-tetramethyl-p-phenylenediamine (TMPD) in a cyanide-sensitive fashion. The ascorbate oxidation was coupled to the uphill Na+ extrusion which was stimulated by CCCP and a penetrating weak base, diethylamine, as well as by valinomycin with or without diethylamine.Operation of the Bacillus FTU terminal oxidase resulted in the generation of a Ay which, in the Na' medium, was slightly decreased by CCCP and strongly decreased by CCCP + diethylamine. In the K' medium, CCCP discharged Ay even without diethylamine. Ascorbate oxidation was competent in ATP synthesis which was resistant to CCCP in the Na' medium and sensitive to CCCP in the K + medium as if Na'-and H+-coupled oxidative phosphorylations were operative in the Na' and K + media, respectively. Inside-out subcellular vesicles of Bacillus FTU were found to be competent in the Na+ uptake supported by oxidation of ascorbate + TMPD or diaminodurene. CCCP or valinomycin + K' increased the Na' uptake very strongly. The process was completely inhibited by cyanide or monensin, the former, but not the latter, being inhibitory for respiration. The data obtained indicate that in Bacillus FTU there is not only H'-motive but also Na+-motive terminal oxidase activity. Numerous studies carried out over the past few years have shown clearly that H + is not unique as a coupling ion. A similar role can, in fact, be performed by Na' (for reviews, see [l -41). In particular, it was found that the sodium potential (ADNa) can be formed by an Na+-motive respiratory chain [5 -81, Na+-ATPase [9 -111 or Na+-dependent decarboxylases [9, 12, 131, localized in the cytoplasmic membrane of some bacteria, and by Na'/K+-ATPase in the plasma membrane of animal cells. The A,&+ formed is utilized for ATP synthesis [9, 141, for uphill transport of many metabolites [15] and for bacterial flagellum rotation [16, 171. Until recently, the only respiratory chain reaction competent in AilNa formation was assumed to be the Na'-motive NADH -quinone reductase found in Vibrio alginolyticus [18], halo-tolerant bacterium Bal

Involvement of a d‐type oxidase in the Na+‐motive respiratory chain of Escherichia coli growing under low Δ\̄gmH+ conditions

“…The same figures show that the protonophore CCCP has to be added for obtaining measurable Na + uptake. As found previously [1,[7][8][9], this effect results from the CCCP-mediated H* efflux which discharges the electric potential produced by the Na*-motive oxidase. The stimulating action of CCCP does not make it possible to explain observed Na ÷ uptake by co-operation of an H÷-motive oxidase and an endogenous Na*/H" antiporter.…”

Sodium Ion Cycle in Bacterial Pathogens: Evidence from Cross-Genome Comparisons

“…An Na ϩ -transporting terminal oxidase has also been found in a representative of the genus Bacillus (8,115,178), later identified as Bacillus halodurans (71) (Table 1) but, rather, tentatively consider them to be H ϩ pumps (Table 2). Because the cytochrome bo-type type terminal oxidase of E. coli has been directly demonstrated to be an H ϩ pump (162,163), similar enzymes in other bacteria are generally assumed to be specific to H ϩ ions.…”

Submicromolar Ag+ increases passive Na+ permeability and inhibits the respiration‐supported formation of Na+ gradient in Bacillus FTU vesicles

“…To obtain ultrasonic inside-out subcellular vesicles, the previously described method [2] was employed.…”