The Experts below are selected from a list of 2007 Experts worldwide ranked by ideXlab platform
Terry A. Krulwich - One of the best experts on this subject based on the ideXlab platform.
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Alkaliphilic bacteria with impact on industrial applications, concepts of early life forms, and bioenergetics of ATP synthesis
Frontiers in bioengineering and biotechnology, 2015Co-Authors: Laura Preiss, David Hicks, Shino Suzuki, Thomas Meier, Terry A. KrulwichAbstract:Alkaliphilic bacteria typically grow well at pH 9, with the most extremophilic strains growing up to pH values as high as pH 12-13. Interest in extreme Alkaliphiles arises because they are sources of useful, stable enzymes, and the cells themselves can be used for biotechnological and other applications at high pH. In addition, alkaline hydrothermal vents represent an early evolutionary niche for Alkaliphiles and novel extreme Alkaliphiles have also recently been found in alkaline serpentinizing sites. A third focus of interest in Alkaliphiles is the challenge raised by the use of proton-coupled ATP synthases for oxidative phosphorylation by non-fermentative Alkaliphiles. This creates a problem with respect to tenets of the chemiosmotic model that remains the core model for the bioenergetics of oxidative phosphorylation. Each of these facets of alkaliphilic bacteria will be discussed with a focus on extremely alkaliphilic Bacillus strains. These alkaliphilic bacteria have provided a cogent experimental system to probe adaptations that enable their growth and oxidative phosphorylation at high pH. Adaptations are clearly needed to enable secreted or partially exposed enzymes or protein complexes to function at the high external pH. Also, Alkaliphiles must maintain a cytoplasmic pH that is significantly lower than the pH of the outside medium. This protects cytoplasmic components from an external pH that is alkaline enough to impair their stability or function. However, the pH gradient across the cytoplasmic membrane, with its orientation of more acidic inside than outside, is in the reverse of the productive orientation for bioenergetic work. The reversed gradient reduces the trans-membrane proton motive force available to energize ATP synthesis. Multiple strategies are hypothesized to be involved in enabling Alkaliphiles to circumvent the challenge of a low bulk proton-motive force energizing proton-coupled ATP synthesis at high pH.
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Bioenergetic Adaptations That Support Alkaliphily
Physiology and Biochemistry of Extremophiles, 2014Co-Authors: Terry A. Krulwich, David Hicks, Talia H. Swartz, Masahiro ItoAbstract:Two themes that run through this chapter are the whole-cell, systems biology aspects of alkaliphile bioenergetics and the diverse ion transporters, pumps, and channels that participate in this system, many of which were first discovered in Alkaliphiles and many of which have alkaliphile-specific roles or adaptations. All Alkaliphiles examined to date, including both anaerobes and aerobes, do indeed maintain a cytoplasmic pH much lower than the external pH. The growing amount of comparative genomic data between Alkaliphiles and neutrophiles has made it much easier to identify putative alkaliphile-specific deviations in conserved and functionally important residues or motifs in proteins of bioenergetic interest. Compelling genomic and biochemical evidence attest to the fact that extreme Alkaliphiles experience a low proton motive force (PMF) at high pH. Alkaliphily in bacteria depends upon one or more Na+/H+ antiporters that catalyze proton uptake in exchange for cytoplasmic Na+. The specific properties of the antiporters of alkaliphilic Bacillus that support its functions are not yet clear, but antiporter properties of interest in relation to alkaliphily have emerged for a different alkaliphile. The proton transfer might involve direct protein–protein interactions with a respiratory chain complex, as suggested by for mitochondria, and/or involve the abundant cardiolipin of the alkaliphile membrane.
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2.6 Adaptive Mechanisms of Extreme Alkaliphiles
2011Co-Authors: Terry A. Krulwich, Masahiro Ito, Jun Liu, Makoto Fujisawa, Masato Morino, David B. HicksAbstract:Prologue . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120 Cytoplasmic pH Homeostasis: The Central Challenge . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121 Growth of Extreme Alkaliphiles at Alkaline Cytoplasmic pH Values Not Tolerated by Neutralophiles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 123 Na/H Antiporters: Key Participants in Cytoplasmic pH Homeostasis of Alkaliphiles . 124 Redundancy in the Cation/Proton Antiporter Complements of Alkaliphiles, and the Importance of Mrp-Type Antiporters in Alkaliphily . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 125 Additional Structural, Enzymatic, and Metabolic Strategies for Cytoplasmic pH Homeostasis, and Their Built-In Redundancies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 130 A Low Bulk Protonmotive Force (pmf) Resulting from Successful pH Homeostasis, and Its Bioenergetic Consequences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 131 Strategies for Oxidative Phosphorylation at Low Protonmotive Force . . . . . . . . . . . . . . . . . . . 132 Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135 Cross-References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135
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The ATP Synthase a-subunit of Extreme Alkaliphiles Is a Distinct Variant MUTATIONS IN THE CRITICAL ALKALIPHILE-SPECIFIC RESIDUE LYS-180 AND OTHER RESIDUES THAT SUPPORT ALKALIPHILE OXIDATIVE PHOSPHORYLATION
The Journal of biological chemistry, 2010Co-Authors: Makoto Fujisawa, Terry A. Krulwich, Jun Liu, Oliver J. Fackelmayer, David HicksAbstract:A lysine residue in the putative proton uptake pathway of the ATP synthase a-subunit is found only in alkaliphilic Bacillus species and is proposed to play roles in proton capture, retention and passage to the synthase rotor. Here, Lys-180 was replaced with alanine (Ala), glycine (Gly), cysteine (Cys), arginine (Arg), or histidine (His) in the chromosome of alkaliphilic Bacillus pseudofirmus OF4. All mutants exhibited octylglucoside-stimulated ATPase activity and β-subunit levels at least as high as wild-type. Purified mutant F1F0-ATP synthases all contained substantial a-subunit levels. The mutants exhibited diverse patterns of native (no octylglucoside) ATPase activity and a range of defects in malate growth and in vitro ATP synthesis at pH 10.5. ATP synthesis by the Ala, Gly, and His mutants was also impaired at pH 7.5 in the presence of a protonophoric uncoupler. Thus Lys-180 plays a role when the protonmotive force is reduced at near neutral, not just at high pH. The Arg mutant exhibited no ATP synthesis activity in the alkaliphile setting although activity was reported for a K180R mutant of a thermoalkaliphile synthase (McMillan, D. G., Keis, S., Dimroth, P., and Cook, G. M. (2007) J. Biol. Chem. 282, 17395–17404). The hypothesis that a-subunits from extreme Alkaliphiles and the thermoalkaliphile represent distinct variants was supported by demonstration of the importance of additional alkaliphile-specific a-subunit residues, not found in the thermoalkaliphile, for malate growth of B. pseudofirmus OF4. Finally, a mutant B. pseudofirmus OF4 synthase with switched positions of Lys-180 (helix 4) and Gly-212 (helix 5) retained significant coupled synthase activity accompanied by proton leakiness.
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F1F0-ATP synthases of alkaliphilic bacteria: lessons from their adaptations
Biochimica et biophysica acta, 2010Co-Authors: David Hicks, Jun Liu, Makoto Fujisawa, Terry A. KrulwichAbstract:Abstract This review focuses on the ATP synthases of alkaliphilic bacteria and, in particular, those that successfully overcome the bioenergetic challenges of achieving robust H + -coupled ATP synthesis at external pH values > 10. At such pH values the protonmotive force, which is posited to provide the energetic driving force for ATP synthesis, is too low to account for the ATP synthesis observed. The protonmotive force is lowered at a very high pH by the need to maintain a cytoplasmic pH well below the pH outside, which results in an energetically adverse pH gradient. Several anticipated solutions to this bioenergetic conundrum have been ruled out. Although the transmembrane sodium motive force is high under alkaline conditions, respiratory alkaliphilic bacteria do not use Na + - instead of H + -coupled ATP synthases. Nor do they offset the adverse pH gradient with a compensatory increase in the transmembrane electrical potential component of the protonmotive force. Moreover, studies of ATP synthase rotors indicate that Alkaliphiles cannot fully resolve the energetic problem by using an ATP synthase with a large number of c -subunits in the synthase rotor ring. Increased attention now focuses on delocalized gradients near the membrane surface and H + transfers to ATP synthases via membrane-associated microcircuits between the H + pumping complexes and synthases. Microcircuits likely depend upon proximity of pumps and synthases, specific membrane properties and specific adaptations of the participating enzyme complexes. ATP synthesis in Alkaliphiles depends upon alkaliphile-specific adaptations of the ATP synthase and there is also evidence for alkaliphile-specific adaptations of respiratory chain components.
Isao Yumoto - One of the best experts on this subject based on the ideXlab platform.
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Contribution of intracellular negative ion capacity to Donnan effect across the membrane in alkaliphilic Bacillus spp.
Journal of bioenergetics and biomembranes, 2016Co-Authors: Toshitaka Goto, Toshinao Hirabayashi, Hajime Morimoto, Koji Yamazaki, Norio Inoue, Hidetoshi Matsuyama, Isao YumotoAbstract:To elucidate the energy production mechanism of Alkaliphiles, the relationship between the H+ extrusion rate by the respiratory chain and the corresponding ATP synthesis rate was determined in the facultative alkaliphile Bacillus cohnii YN-2000 and compared with those in the obligate alkaliphile Bacillus clarkii DSM 8720T and the neutralophile Bacillus subtilis IAM 1026. Under high aeration condition, much higher ATP synthesis rates and larger Δψ in the alkaliphilic Bacillus spp. grown at pH 10 than those in the neutralophilic B. subtilis grown at pH 7 were observed. This high ATP productivity could be attributed to the larger Δψ in Alkaliphiles than in B. subtilis because the H+ extrusion rate in Alkaliphiles cannot account for the high ATP productivity. However, the large Δψ in the Alkaliphiles could not be explained only by the H+ translocation rate in the respiratory chain in Alkaliphiles. There is a possibility that the Donnan effect across the membrane has the potential to contribute to the large Δψ. To estimate the contribution of the Donnan effect to the large Δψ in alkaliphilic Bacillus spp. grown at pH 10, intracellular negative ion capacity was examined. The intracellular negative ion capacities in Alkaliphiles grown at pH 10 under high aeration condition corresponding to their intracellular pH (pH 8.1) were much higher than those in Alkaliphiles grown under low aeration condition. A proportional relationship is revealed between the negative ion capacity and Δψ in Alkaliphiles grown under different aeration conditions. This relationship strongly suggests that the intracellular negative ion capacity contributes to the formation of Δψ through the Donnan effect in alkaliphilic Bacillus spp. grown at pH 10.
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Environmental and Taxonomic Biodiversities of Gram-Positive Alkaliphiles
Physiology and Biochemistry of Extremophiles, 2014Co-Authors: Isao YumotoAbstract:This chapter focuses on the environmental and taxonomic distributions of gram-positive Alkaliphiles. Garbeva et al. developed a polymerase chain reaction (PCR) system for studying the diversity of the species of Bacillus and related taxa using DNA directly obtained from soil. Detection of Bacillus halodurans by this procedure indicated that although the soil samples were slightly acidic, Bacillus halodurans might be one of the major Bacillus species in the soil samples used in that study. Microbial diversities of soda lakes in Africa, Europe, and North America have been detected on the basis of the analysis of DNA clone libraries produced by amplification of obtained DNA as well as from the isolation of microorganisms from the environments. The major gram-negative isolates are members of the gamma subdivision of Proteobacteria. Indigo-reducing bacteria have been isolated by Takahara and Tanabe and identified as Bacillus sp. they have been named Bacillus alcalophilus. This is the only species that can grow at 5°C among the currently known alkaliphilic Bacillus spp. The chapter provides facts that suggest that niches of Bacillus patagoniensis are in soil and in rhizosphere of certain plants. Some of the strains in this group were formally classified as Bacillus. In the next decade, the understanding of the distribution in the environment and of the taxonomic diversities of Alkaliphiles will proceed further not only by isolation of novel species of Alkaliphiles but also from results of analyses of DNA directly obtained from various environments.
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2 3 environmental distribution and taxonomic diversity of Alkaliphiles
2010Co-Authors: Koki Horikoshi, Isao Yumoto, Kazuaki YoshimuneAbstract:Background . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 56 Aims and Significance of Taxonomy of Alkaliphilic Fermicutes . . . . . . . . . . . . . . . . . . . . . . . . . . . 57 Environmental Distributions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59 Soil Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59 Soda Lakes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61 Gut of Insects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64 Groundwater . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65 Sea and Sea-Related Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65 Indigo Fermentation Liquor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66 Other Environments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67 Taxonomy of Isolated Alkaliphilic Firmicutes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68 Genus Bacillus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68 Conclusions and Perspective . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 75 Cross-References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 76
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Bacillus polygoni sp. nov., a moderately halophilic, non-motile obligate alkaliphile isolated from indigo balls.
INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, 2008Co-Authors: Kenichi Aino, Hidetoshi Matsuyama, Toshihide Matsuno, Kikue Hirota, Naoki Morita, Yoshinobu Nodasaka, Taketomo Fujiwara, Kazuaki Yoshimune, Isao YumotoAbstract:A moderately halophilic, obligate alkaliphile (growth range pH 8–12), designated strain YN-1T, was isolated from indigo balls obtained from Ibaraki, Japan. The cells of the isolate stained Gram-positive, and were aerobic, non-motile, sporulating rods which grew optimally at pH 9. The strain grew in 3–14 % NaCl with optimum growth in 5 % NaCl. It hydrolysed casein and Tweens 20, 40 and 60, but not gelatin, starch, DNA or pullulan. Its major isoprenoid quinone was MK-7 and its cellular fatty acid profile mainly consisted of anteiso-C15 : 0, anteiso-C17 : 0 and anteiso-C17 : 1. 16S rRNA phylogeny suggested that strain YN-1T was a member of group 7 (Alkaliphiles) of the genus Bacillus, with the closest relative being Bacillus clarkii DSM 8720T (similarity 99.5 %). However, DNA–DNA hybridization showed a low DNA–DNA relatedness (7 %) of strain YN-1T with B. clarkii DSM 8720T. Owing to the significant differences in phenotypic and chemotaxonomic characteristics, and phylogenetic and DNA–DNA relatedness data, the isolate merits classification as a new species, for which the name Bacillus polygoni is proposed. The type strain of this species is YN-1T (=JCM 14604T=NCIMB 14282T).
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Cytochrome c and bioenergetic hypothetical model for alkaliphilic Bacillus spp.
Journal of bioscience and bioengineering, 2005Co-Authors: Toshitaka Goto, Koji Yamazaki, Norio Inoue, Hidetoshi Matsuyama, Toshihide Matsuno, Megumi Hishinuma-narisawa, Isao YumotoAbstract:Although a bioenergetic parameter is unfavorable for production of ATP (ΔpH Bacillus strains are higher than those of neutralophilic Bacillus subtilis . This finding suggests that Alkaliphiles possess a unique energy-producing machinery taking advantage of the alkaline environment. Expected bioenergetic parameters for the production of ATP (ΔpH and ΔΨ) do not reflect the actual parameters for energy production. Certain strains of alkaliphilic Bacillus spp. possess large amounts of cytochrome c when grown at a high pH. The growth rate and yield are higher at pH 10 than at pH 7 in facultative Alkaliphiles. These findings suggest that a large amount of cytochrome c at high pHs ( e.g. , pH 10) may be advantageous for sustaining growth. To date, isolated cytochromes c of Alkaliphiles have a very low midpoint redox potential (less than +100 mV) compared with those of neutralophiles (approximately +220 mV). On the other hand, the redox potential of the electron acceptor from cytochrome c , that is, cytochrome c oxidase, seems to be normal (redox potential of cytochrome a =+250 mV). This large difference in midpoint redox potential between cytochrome c and cytochrome a concomitant with the configuration ( e.g. , a larger negative ion capacity at the inner surface membrane than at the outer surface for the attraction of H + to the intracellular membrane and a large amount of cyrochrome c ) supporting H + -coupled electron transfer of cytochrome c may have an important meaning in the adaptation of Alkaliphiles at high pHs. This respiratory system includes a more rapid and efficient H + and e − flow across the membrane in Alkaliphiles than in neutralophiles.
Gerard Muyzer - One of the best experts on this subject based on the ideXlab platform.
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of reductive sulfur cycle from soda lakes
2015Co-Authors: Gerard MuyzerAbstract:Abstract Anaerobic enrichments with H2 as electron donor and thiosulfate/polysulfide as electron acceptor at pH 10 and 0.6 M total Na+ yielded two non sulfate-reducing representatives of reductive sulfur cycle from soda lake sediments. Strain AHT 1 was isolated with thiosulfate as the electron acceptor from north–eastern Mongolian soda lakes and strain AHT 2—with polysulfide as the electron acceptor from Wadi al Natrun lakes in Egypt. Both isolates represented new phylogenetic lineages: AHT 1—within Clostridiales and AHT 2—within the Deltaproteobacteria. Both bacteria are obligate anaerobes with respiratory metabolism. Both grew chemolithoautotrophically with H2 as the electron donor and can use thiosulfate, elemental sulfur and polysulfide as the electron acceptors. AHT 2 also used nitrate as acceptor, reducing it to ammonia. During thiosulfate reduction, AHT 1 excreted sulfite. dsrAB gene was not found in either strain. Both strains were moderate salt-tolerant (grow up to 2 M total Na+) true Alkaliphiles (grow between pH 8.5 and 10.3). On the basis of the phenotypic and phylogenetic data, strains AHT 1 and AHT 2 are proposed as new genera and species Dethiobacter alkaliphilus and Desulfurivibrio alkaliphilus, respectively
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dethiobacter alkaliphilus gen nov sp nov and desulfurivibrio alkaliphilus gen nov sp nov two novel representatives of reductive sulfur cycle from soda lakes
Extremophiles, 2008Co-Authors: D. Y. Sorokin, T P Tourova, M Mussmann, Gerard MuyzerAbstract:Anaerobic enrichments with H2 as electron donor and thiosulfate/polysulfide as electron acceptor at pH 10 and 0.6 M total Na+ yielded two non sulfate-reducing representatives of reductive sulfur cycle from soda lake sediments. Strain AHT 1 was isolated with thiosulfate as the electron acceptor from north–eastern Mongolian soda lakes and strain AHT 2—with polysulfide as the electron acceptor from Wadi al Natrun lakes in Egypt. Both isolates represented new phylogenetic lineages: AHT 1—within Clostridiales and AHT 2—within the Deltaproteobacteria. Both bacteria are obligate anaerobes with respiratory metabolism. Both grew chemolithoautotrophically with H2 as the electron donor and can use thiosulfate, elemental sulfur and polysulfide as the electron acceptors. AHT 2 also used nitrate as acceptor, reducing it to ammonia. During thiosulfate reduction, AHT 1 excreted sulfite. dsrAB gene was not found in either strain. Both strains were moderate salt-tolerant (grow up to 2 M total Na+) true Alkaliphiles (grow between pH 8.5 and 10.3). On the basis of the phenotypic and phylogenetic data, strains AHT 1 and AHT 2 are proposed as new genera and species Dethiobacter alkaliphilus and Desulfurivibrio alkaliphilus, respectively.
David Hicks - One of the best experts on this subject based on the ideXlab platform.
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Alkaliphilic bacteria with impact on industrial applications, concepts of early life forms, and bioenergetics of ATP synthesis
Frontiers in bioengineering and biotechnology, 2015Co-Authors: Laura Preiss, David Hicks, Shino Suzuki, Thomas Meier, Terry A. KrulwichAbstract:Alkaliphilic bacteria typically grow well at pH 9, with the most extremophilic strains growing up to pH values as high as pH 12-13. Interest in extreme Alkaliphiles arises because they are sources of useful, stable enzymes, and the cells themselves can be used for biotechnological and other applications at high pH. In addition, alkaline hydrothermal vents represent an early evolutionary niche for Alkaliphiles and novel extreme Alkaliphiles have also recently been found in alkaline serpentinizing sites. A third focus of interest in Alkaliphiles is the challenge raised by the use of proton-coupled ATP synthases for oxidative phosphorylation by non-fermentative Alkaliphiles. This creates a problem with respect to tenets of the chemiosmotic model that remains the core model for the bioenergetics of oxidative phosphorylation. Each of these facets of alkaliphilic bacteria will be discussed with a focus on extremely alkaliphilic Bacillus strains. These alkaliphilic bacteria have provided a cogent experimental system to probe adaptations that enable their growth and oxidative phosphorylation at high pH. Adaptations are clearly needed to enable secreted or partially exposed enzymes or protein complexes to function at the high external pH. Also, Alkaliphiles must maintain a cytoplasmic pH that is significantly lower than the pH of the outside medium. This protects cytoplasmic components from an external pH that is alkaline enough to impair their stability or function. However, the pH gradient across the cytoplasmic membrane, with its orientation of more acidic inside than outside, is in the reverse of the productive orientation for bioenergetic work. The reversed gradient reduces the trans-membrane proton motive force available to energize ATP synthesis. Multiple strategies are hypothesized to be involved in enabling Alkaliphiles to circumvent the challenge of a low bulk proton-motive force energizing proton-coupled ATP synthesis at high pH.
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Bioenergetic Adaptations That Support Alkaliphily
Physiology and Biochemistry of Extremophiles, 2014Co-Authors: Terry A. Krulwich, David Hicks, Talia H. Swartz, Masahiro ItoAbstract:Two themes that run through this chapter are the whole-cell, systems biology aspects of alkaliphile bioenergetics and the diverse ion transporters, pumps, and channels that participate in this system, many of which were first discovered in Alkaliphiles and many of which have alkaliphile-specific roles or adaptations. All Alkaliphiles examined to date, including both anaerobes and aerobes, do indeed maintain a cytoplasmic pH much lower than the external pH. The growing amount of comparative genomic data between Alkaliphiles and neutrophiles has made it much easier to identify putative alkaliphile-specific deviations in conserved and functionally important residues or motifs in proteins of bioenergetic interest. Compelling genomic and biochemical evidence attest to the fact that extreme Alkaliphiles experience a low proton motive force (PMF) at high pH. Alkaliphily in bacteria depends upon one or more Na+/H+ antiporters that catalyze proton uptake in exchange for cytoplasmic Na+. The specific properties of the antiporters of alkaliphilic Bacillus that support its functions are not yet clear, but antiporter properties of interest in relation to alkaliphily have emerged for a different alkaliphile. The proton transfer might involve direct protein–protein interactions with a respiratory chain complex, as suggested by for mitochondria, and/or involve the abundant cardiolipin of the alkaliphile membrane.
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Genome of alkaliphilic Bacillus pseudofirmus OF4 reveals adaptations that support the ability to grow in an external pH range from 7.5 to 11.4.
Environmental microbiology, 2011Co-Authors: Benjamin Janto, Masahiro Ito, David Hicks, Jun Liu, Oliver J. Fackelmayer, Azad Ahmed, Sarah Pagni, Terry Ann Smith, Joshua P. Earl, Liam D. H. ElbourneAbstract:Summary Bacillus pseudofirmus OF4 is an extreme but facultative alkaliphile that grows non-fermentatively in a pH range from 7.5 to above 11.4 and can withstand large sudden increases in external pH. It is a model organism for studies of bioenergetics at high pH, at which energy demands are higher than at neutral pH because both cytoplasmic pH homeostasis and ATP synthesis require more energy. The alkaliphile also tolerates a cytoplasmic pH > 9.0 at external pH values at which the pH homeostasis capacity is exceeded, and manages other stresses that are exacerbated at alkaline pH, e.g. sodium, oxidative and cell wall stresses. The genome of B. pseudofirmus OF4 includes two plasmids that are lost from some mutants without viability loss. The plasmids may provide a reservoir of mobile elements that promote adaptive chromosomal rearrangements under particular environmental conditions. The genome also reveals a more acidic pI profile for proteins exposed on the outer surface than found in neutralophiles. A large array of transporters and regulatory genes are predicted to protect the alkaliphile from its overlapping stresses. In addition, unanticipated metabolic versatility was observed, which could ensure requisite energy for alkaliphily under diverse conditions.
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The ATP Synthase a-subunit of Extreme Alkaliphiles Is a Distinct Variant MUTATIONS IN THE CRITICAL ALKALIPHILE-SPECIFIC RESIDUE LYS-180 AND OTHER RESIDUES THAT SUPPORT ALKALIPHILE OXIDATIVE PHOSPHORYLATION
The Journal of biological chemistry, 2010Co-Authors: Makoto Fujisawa, Terry A. Krulwich, Jun Liu, Oliver J. Fackelmayer, David HicksAbstract:A lysine residue in the putative proton uptake pathway of the ATP synthase a-subunit is found only in alkaliphilic Bacillus species and is proposed to play roles in proton capture, retention and passage to the synthase rotor. Here, Lys-180 was replaced with alanine (Ala), glycine (Gly), cysteine (Cys), arginine (Arg), or histidine (His) in the chromosome of alkaliphilic Bacillus pseudofirmus OF4. All mutants exhibited octylglucoside-stimulated ATPase activity and β-subunit levels at least as high as wild-type. Purified mutant F1F0-ATP synthases all contained substantial a-subunit levels. The mutants exhibited diverse patterns of native (no octylglucoside) ATPase activity and a range of defects in malate growth and in vitro ATP synthesis at pH 10.5. ATP synthesis by the Ala, Gly, and His mutants was also impaired at pH 7.5 in the presence of a protonophoric uncoupler. Thus Lys-180 plays a role when the protonmotive force is reduced at near neutral, not just at high pH. The Arg mutant exhibited no ATP synthesis activity in the alkaliphile setting although activity was reported for a K180R mutant of a thermoalkaliphile synthase (McMillan, D. G., Keis, S., Dimroth, P., and Cook, G. M. (2007) J. Biol. Chem. 282, 17395–17404). The hypothesis that a-subunits from extreme Alkaliphiles and the thermoalkaliphile represent distinct variants was supported by demonstration of the importance of additional alkaliphile-specific a-subunit residues, not found in the thermoalkaliphile, for malate growth of B. pseudofirmus OF4. Finally, a mutant B. pseudofirmus OF4 synthase with switched positions of Lys-180 (helix 4) and Gly-212 (helix 5) retained significant coupled synthase activity accompanied by proton leakiness.
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F1F0-ATP synthases of alkaliphilic bacteria: lessons from their adaptations
Biochimica et biophysica acta, 2010Co-Authors: David Hicks, Jun Liu, Makoto Fujisawa, Terry A. KrulwichAbstract:Abstract This review focuses on the ATP synthases of alkaliphilic bacteria and, in particular, those that successfully overcome the bioenergetic challenges of achieving robust H + -coupled ATP synthesis at external pH values > 10. At such pH values the protonmotive force, which is posited to provide the energetic driving force for ATP synthesis, is too low to account for the ATP synthesis observed. The protonmotive force is lowered at a very high pH by the need to maintain a cytoplasmic pH well below the pH outside, which results in an energetically adverse pH gradient. Several anticipated solutions to this bioenergetic conundrum have been ruled out. Although the transmembrane sodium motive force is high under alkaline conditions, respiratory alkaliphilic bacteria do not use Na + - instead of H + -coupled ATP synthases. Nor do they offset the adverse pH gradient with a compensatory increase in the transmembrane electrical potential component of the protonmotive force. Moreover, studies of ATP synthase rotors indicate that Alkaliphiles cannot fully resolve the energetic problem by using an ATP synthase with a large number of c -subunits in the synthase rotor ring. Increased attention now focuses on delocalized gradients near the membrane surface and H + transfers to ATP synthases via membrane-associated microcircuits between the H + pumping complexes and synthases. Microcircuits likely depend upon proximity of pumps and synthases, specific membrane properties and specific adaptations of the participating enzyme complexes. ATP synthesis in Alkaliphiles depends upon alkaliphile-specific adaptations of the ATP synthase and there is also evidence for alkaliphile-specific adaptations of respiratory chain components.
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Alkaliphilus serpentinus sp. nov. and Alkaliphilus pronyensis sp. nov., two novel anaerobic alkaliphilic species isolated from the serpentinite-hosted Prony Bay Hydrothermal Field (New Caledonia)
Systematic and Applied Microbiology, 2021Co-Authors: Anne Postec, Marianne Quemeneur, Aurélien Lecoeuvre, Nicolas Chabert, Manon Joseph, Gaël ErausoAbstract:Two novel anaerobic alkaliphilic strains, designated as LacT T and LacV T , were isolated from the Prony Bay Hydrothermal Field (PBHF, New Caledonia). Cells were motile, Gram-positive, terminal endosporeforming rods, displaying a straight to curved morphology during the exponential phase. Strains LacT T and LacV T were mesophilic (optimum 30°C), moderately alkaliphilic (optimum pH 8.2 and 8.7, respectively) and halotolerant (optimum 2% and 2.5% NaCl, respectively). Both strains were able to ferment yeast extract, peptone and casamino acids, but only strain LacT T could use sugars (glucose, maltose and sucrose). Both strains disproportionated crotonate into acetate and butyrate. Phylogenetic analysis revealed that strains LacT T and LacV T shared 96.4% 16S rRNA gene sequence identity and were most closely related to A. peptidifermentans Z-7036, A. namsaraevii X-07-2 and A. hydrothermalis FatMR1 (95.7%-96.3%). Their genome size was of 3.29 Mb for strain LacT T and 3.06 Mb for strain LacV T with a G + C content of 36.0 and 33.9 mol%, respectively. The ANI value between both strains was 73.2 %. Finally, strains LacT T (=DSM 100337 = JCM 30643) and LacV T (=DSM 100017 = JCM 30644) are proposed as two novel species of the genus Alkaliphilus, order Clostridiales, phylum Firmicutes, Alkaliphilus serpentinus sp. nov. and Alkaliphilus pronyensis sp. nov., respectively. The genomes of the three Alkaliphilus species isolated from PBHF were consistently detected in the PBHF chimney metagenomes, although at very low abundance, but not significantly in the metagenomes of other serpentinizing systems (marine or terrestrial) worldwide, suggesting they represent indigenous members of the PBHF microbial ecosystem.
