Alkaline pH homeostasis in bacteria: new insights.

Of all the molecular determinants for growth, the hydronium and hydroxide ions are found naturally in the widest concentration range, from acid mine drainage below pH 0 to soda lakes above pH 13. Most bacteria and archaea have mechanisms that maintain their internal, cytoplasmic pH within a narrower range than the pH outside the cell, termed "pH homeostasis." Some mechanisms of pH homeostasis are specific to particular species or groups of microorganisms while some common principles apply across the pH spectrum. The measurement of internal pH of microbes presents challenges, which are addressed by a range of techniques under varying growth conditions. This review compares and contrasts cytoplasmic pH homeostasis in acidophilic, neutralophilic, and alkaliphilic bacteria and archaea under conditions of growth, non-growth survival, and biofilms. We present diverse mechanisms of pH homeostasis including cell buffering, adaptations of membrane structure, active ion transport, and metabolic consumption of acids and bases.

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Cytoplasmic pH Measurement and Homeostasis in Bacteria and Archaea

Na(+)/H(+) antiporters.

Na a /H a antiporters are membrane proteins that play a major role in pH and Na a homeostasis of cells throughout the biological kingdom, from bacteria to humans and higher plants. The emerging genomic sequence projects already have started to reveal that the Na a /H a antiporters cluster in several families. Structure and function studies of a purified antiporter protein have as yet been conducted mainly with NhaA, the key Na a /H a antiporter of Escherichia coli. This antiporter has been overexpressed, purified and reconstituted in a functional form in proteoliposomes. It has recently been crystallized in both 3D as well as 2D crystals. The NhaA 2D crystals were analyzed by cryoelectron microscopy and a density map at 4 A O resolution was obtained and a 3D map was reconstructed. NhaA is shown to exist in the 2D crystals as a dimer of monomers each composed of 12 transmembrane segments with an asymmetric helix packing. This is the first insight into the structure of a polytopic membrane protein. Many Na a /H a antiporters are characterized by very dramatic sensitivity to pH, a property that corroborates their role in pH homeostasis. The molecular mechanism underlying this pH sensitivity has been studied in NhaA. Amino acid residues involved in the pH response have been identified. Conformational changes transducing the pH change into a change in activity were found in loop VIII^IX and at the N-terminus by probing trypsin digestion or binding of a specific monoclonal antibody respectively. Regulation by pH of the eukaryotic Na a /H a antiporters involves an intricate signal transduction pathway (recently reviewed by Yun et al., Am. J. Physiol. 269 (1995) G1^G11). The transcription of NhaA has been shown to be regulated by a novel Na a -specific regulatory network. It is envisaged that interdisciplinary approaches combining structure, molecular and cell biology as well as genomics should be applied in the future to the study of this important group of transporters. fl 2001 Elsevier Science B.V. All rights reserved.

Na á /H á antiporters

All eukaryotic cells contain a wide variety of proteins embedded in the plasma and internal membranes, which ensure transmembrane solute transport. It is now established that a large proportion of these transport proteins can be grouped into families apparently conserved throughout organisms. This article presents the data of an in silicio analysis aimed at establishing a preliminary classification of membrane transport proteins in Saccharomyces cerevisiae. This analysis was conducted at a time when about 65% of all yeast genes were available in public databases. In addition to approximately 60 transport proteins whose function was at least partially known, approximately 100 deduced protein sequences of unknown function display significant sequence similarity to membrane transport proteins characterized in yeast and/or other organisms. While some protein families have been well characterized by classical genetic experimental approaches, others have largely if not totally escaped characterization. The proteins revealed by this in silicio analysis also include a putative K+ channel, proteins similar to aquaporins of plant and animal origin, proteins similar to Na+-solute symporters, a protein very similar to electroneural cation-chloride cotransporters, and a putative Na+-H+ antiporter. A new research area is anticipated: the functional analysis of many transport proteins whose existence was revealed by genome sequencing.

An overview of membrane transport proteins in Saccharomyces cerevisiae.

The thermophilic, peptidolytic, anaerobic bacterium Clostridium fervidus is unable to generate a pH gradient in the range of 5.5-8.0, which limits growth of the organism to a narrow pH range (6.3-7.7). A significant membrane potential (delta psi approximately -60 mV) and chemical gradient of Na+ (-Z delta pNa approximately -60 mV) are formed in the presence of metabolizable substrates. Energy-dependent Na+ efflux is inhibited by the Na+/H+ ionophore monensin but is stimulated by uncouplers, suggesting that the Na+ gradient is formed by a primary pumping mechanism rather than by secondary Na+/H+ antiport. This primary sodium pump was found to be an ATPase that has been characterized in inside-out membrane vesicles and in proteoliposomes in which solubilized ATPase was reconstituted. The enzyme is stimulated by Na+, resistant to vanadate, and sensitive to nitrate, which is indicative of an F/V-type Na(+)-ATPase. In the proteoliposomes Na+ accumulation depends on the presence of ATP, is inhibited by the ATPase inhibitor nitrate, and is completely prevented by the ionophore monensin but is stimulated by protonophores and valinomycin. These and previous observations, which indicated that secondary amino acid transport uses solely Na+ as coupling ion, demonstrate that energy transduction at the membrane in C. fervidus is exclusively dependent on a Na+ cycle.

/pdf/energy-transduction-in-the-thermophilic-anaerobic-bacterium-3krsi2hjn5.pdf

Energy transduction in the thermophilic anaerobic bacterium Clostridium fervidus is exclusively coupled to sodium ions.

https://pure.rug.nl/ws/files/3285018/1994BiochimBiophysActaLolkema.pdf

Na +-coupled versus H +-coupled energy transduction in bacteria

https://pure.rug.nl/ws/files/3302289/1993BiochimBiophysActaSpeelmans.pdf

Gea Speelmans

Papers

Energy transduction in the thermophilic anaerobic bacterium Clostridium fervidus is exclusively coupled to sodium ions.

Na +-coupled versus H +-coupled energy transduction in bacteria

Application of thermostable reaction centers from Chloroflexus aurantiacus as a protonmotive force generating system

Sodium coupled transport in bacteria

AminoAcidTransport intheThermophilic Anaerobe Clostridium fervidus IsDriven byan Electrochemical SodiumGradient