Fractionation of sulfur isotopes during dissimilatory reduction of sulfate by a thermophilic gram-negative bacterium at 60 °C

Sulfur isotope (³⁴S/³²S) fractionation during reduction of dissolved sulfate was investigated with a growing batch culture of a thermophilic, gram-negative, sulfate-reducing bacterium (strain MT-96) at 60 °C. The completely oxidizing strain was isolated from geothermally heated sediments of a shallow-water hydrothermal vent in the Mediterranean Sea. The hydrogen sulfide produced in the experiments was enriched in ³²S by approximately 19‰ as compared to sulfate, which indicates that stable isotope discrimination by this thermophile is within the range found previously for mesophilic sulfate-reducing bacteria, and only slightly higher than that observed for the thermophilic gram-positive Desulfotomaculum nigrificans. Received: 1 December 1998 / Accepted: 25 May 1999     

Dissimilatory hexaheme c nitrite reductase of “Spirillum” strain 5175: purification and properties

When grown with nitrate as terminal electron acceptor both the soluble (periplasm, cytoplasm) and the membrane fraction of Spirillum strain 5175 exhibited high nitrite reductase activity. The nitrite reductase obtained from the soluble fraction was purified 76-fold to electrophoretical homogeneity. The enzyme reduced nitrite to ammonia with a specific activity of 723 mol NO _inf2 ^sup- × (mg protein × min)^-1. The molecular mass was 58±1 kDa by SDS-PAGE compared to 59±2 kDa determined by size exclusion chromatography under nondenaturing conditions. The enzyme (as isolated) contained 5.97±0.15 heme c molecules/M_r 58 kDa. The absorption spectrum was typical for c-type cytochrome with maxima at 280, 408, 532 and 610 nm (oxidized) and at 420, 523 and 553 nm (dithionite-reduced). The enzyme (as isolated) exhibited a complex set of high-spin and lowspin ferric heme resonances with g-values at 9.82, 3,85, 3.31, 2.95, 2.30 and 1.49 in agreement with data reported for electron paramagnetic resonance spectra of nitrite reductases from Desulfovibrio desulfuricans, Wolinella succinogenes and Escherichia coli.Abbreviations DNRA dissimilatory nitrate reduction to ammonia - EPR electron paramagnetic resonance - PAGE polyacrylamide gel electrophoresis - NaP_i sodium phosphate - SDS sodium dodecylsulfate     

The influence of seasonal changes on energy metabolism in Mytilus edulise (L.).—III. Anaerobic energy metabolism

• 1.1. Seasonal changes in the accumulation of end products after 48 hr of exposure to air and in the composition of the free amino acid pool were studied in Mytilus edulis.
• 2.2. The accumulation levels of succinate and acetate showed only weak seasonal changes.
• 3.3. Conversion of succinate to propionate was high in summer and virtually zero in winter
• 4.4. Alanine and most other free amino acids were present in relatively high concentrations in summer and early autumn and reached minimal values in winter and early spring.
• 5.5. Exceptions were glutamate, aspartate and taurine, which showed hardly an season related changes and glycine, which changed inversely to the majority of the free amino acids.
• 6.6. The anaerobic formation of alanine was inversely proportional to the endogenous concentration.
• 7.7. The only other free amino acids affected by anaerobiosis were glutamate and aspartate, which respectively increased and decreased under these conditions.

     

Comparative systematic study on “Spirillum” 5175, Campylobacter and Wolinella species

Physiological tests, redetermination of G+C values with HPLC and DNA-DNA hybridization were used to determine the taxonomic affiliation of Spirillum 5175. This facultatively sulfur-reducing bacterium was compared to the type strains of the phenotypically most similar species Wolinella succinogenes and Campylobacter sputorum biovar bubulus. In addition to morphology, the following physiological properties were in common: use of elemental sulfur, nitrate, nitrite, aspartate, fumarate or malate as electron acceptor for growth with hydrogen or formate under anoxic conditions; microaerobic growth with 2% (v/v) oxygen. The G+C content of Wolinella succinogenes (51.8 mol%) and Campylobacter sputorum biovar bubulus (30.4 mol%) differs about 10 mol% from the G+C content of Spirillum 5175 (40.6 mol%). No significant DNA homology could be detected between the three strains. These differences excluded affiliation of Spirillum 5175 with the genera Wolinella or Campylobacter despite phenotypic similarities. On the basis of our results and DNA-rRNA hybridization studies by other authors, we established the new genus Sulfurospirillum for the freeliving Campylobacter-like bacteria Spirillum 5175 and Campylobacter spec. DSM 806. Strain Spirillum 5175 is described as the type strain of the new genus and species Sulfurospirillum deleyianum.Dedicated to R. S. Wolfe on the occasion of his 70th birthday     

Physiological Characterization of an Anaerobic Ammonium-Oxidizing Bacterium Belonging to the “Candidatus Scalindua” Group

The phylogenetic affiliation and physiological characteristics (e.g., K_s and maximum specific growth rate [μ_max]) of an anaerobic ammonium oxidation (anammox) bacterium, “Candidatus Scalindua sp.,” enriched from the marine sediment of Hiroshima Bay, Japan, were investigated. “Candidatus Scalindua sp.” exhibits higher affinity for nitrite and a lower growth rate and yield than the known anammox species.     

Anaerobic metabolism in sublittoral living Mytilus galloprovincialis in the mediterranean—IV. Role of amino acids in adaptation to low salinities during anaerobiosis and aerobiosis

• 1.1. When Mytilus galloprovincialis were transferred from 38 to 19%. sea water (S), the metabolism became anaerobic for at least 8 hr. After 24 hr the animals were entirely aerobic again.
• 2.2. Upon transfer to 19%. S, the total free amino acid concentration in haemolymph doubled within 4 hr, remaining nearly constant thereafter, up to 48 hr.
• 3.3. In the posterior adductor muscle a strong decrease of alanine and glycine occurred at 48 hr exposure to 19%. S, and a smaller decrease of glutamate; taurine remained relatively constant. When transferred again to 38%. S after 14 days, a strong overcompensation occurred in the concentrations of alanine and proline, and a smaller overcompensation in those of threonine and serine.
• 4.4. In the gill no distinct change in the amino acid pool occurred during 14 days of exposure, with the exception of a decrease in serine. When transferred again to 38%. S, a strong overcompensation occurred in alanine, proline, glycine and serine, and a smaller in glutamate and threonine.
• 5.5. No evidence for anaerobic metabolism in the decrease of the amino acid pool was found.
• 6.6. M. galloprovincialis is less able to adapt to low salinities than the more euryhaline M. edulis.

     

“Pinching” the ammonia tunnel of CTP synthase unveils coordinated catalytic and allosteric-dependent control of ammonia passage

Molecular gates within enzymes often play important roles in synchronizing catalytic events. We explored the role of a gate in cytidine-5′-triphosphate synthase (CTPS) from Escherichia coli. This glutamine amidotransferase catalyzes the biosynthesis of CTP from UTP using either l-glutamine or exogenous NH₃ as a substrate. Glutamine is hydrolyzed in the glutaminase domain, with GTP acting as a positive allosteric effector, and the nascent NH₃ passes through a gate located at the end of a ~25-Å tunnel before entering the synthase domain where CTP is generated. Substitution of the gate residue Val 60 by Ala, Cys, Asp, Trp, or Phe using site-directed mutagenesis and subsequent kinetic analyses revealed that V60-substitution impacts glutaminase activity, nucleotide binding, salt-dependent inhibition, and inter-domain NH₃ transport. Surprisingly, the increase in steric bulk present in V60F perturbed the local structure consistent with “pinching” the tunnel, thereby revealing processes that synchronize the transfer of NH₃ from the glutaminase domain to the synthase domain. V60F had a slightly reduced coupling efficiency at maximal glutaminase activity that was ameliorated by slowing down the glutamine hydrolysis reaction, consistent with a “bottleneck” effect. The inability of V60F to use exogenous NH₃ was overcome in the presence of GTP, and more so if CTPS was covalently modified by 6-diazo-5-oxo-l-norleucine. Use of NH₂OH by V60F as an alternative bulkier substrate occurred most efficiently when it was concomitant with the glutaminase reaction. Thus, the glutaminase activity and GTP-dependent activation act in concert to open the NH₃ gate of CTPS to mediate inter-domain NH₃ transport.     

Differential hepatic protein tyrosine nitration of mouse due to aging – Effect on mitochondrial energy metabolism,quality control machinery of the endoplasmic reticulum and metabolism of drugs

Aging is the inevitable fate of life which leads to the gradual loss of functions of different organs and organelles of all living organisms. The liver is no exception. Oxidative damage to proteins and other macromolecules is widely believed to be the primary cause of aging. One form of oxidative damage is tyrosine nitration of proteins, resulting in the potential loss of their functions. In this study, the effect of age on the nitration of tyrosine in mouse liver proteins was examined. Liver proteins from young (19–22 weeks) and old (24 months) C57/BL6 male mice were separated using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS–PAGE) and electroblotted onto nitrocellulose membranes. Proteins undergoing tyrosine nitration were identified using anti-nitrotyrosine antibody. Three different protein bands were found to contain significantly increased levels of nitrotyrosine in old mice (Wilconxon rank-sum test, p < 0.05). Electrospray ionization liquid chromatography tandem mass spectrometry (ESI-LC–MS/MS) was used to identify the proteins in these bands, which included aldehyde dehydrogenase 2, Aldehyde dehydrogenase family 1, subfamily A1, ATP synthase, H⁺ transporting, mitochondrial F1 complex, β subunit, selenium-binding protein 2, and protein disulfide-isomerase precursor. The possible impairment of their functions can lead to altered hepatic activity and have been discussed.     

The dynamics of the diversity–energy relationship during the last 21,000 years

Aim

Spatial diversity patterns are linked to energy availability, but how the diversity–energy relationship changes in space and time is unclear. There are three possible scenarios: (a) equilibrium dynamics, where diversity is always positively related to energy availability in both time and space; (b) out-of-equilibrium dynamics, where diversity is determined by energy availability in equilibrium, but diversity variation lags behind changes in energy availability, leading to a mismatch between temporal and spatial diversity–energy relationships; and (c) disequilibrium dynamics where the equilibrium does not exist or is irrelevant. We attempt to distinguish these scenarios using spatio-temporal palaeoecological data.

Location

USA and Canada.

Time period

21,000–1,000 years before present.

Major taxa studied

Woody plants.

Methods

We tested for the strength and direction of temporal and spatial relationships between pollen type diversity of woody species and energy availability (estimated from temperature and precipitation) in 1,000-year time steps.

Results

Whereas the temporal diversity–energy relationship appears only when energy levels rapidly change, the spatial diversity–energy relationship is pronounced only when these levels stabilize.

Main conclusions

Our findings are consistent with the out-of-equilibrium scenario – diversity equilibria do exist and act as attractors of diversity dynamics, but diversity is often out of equilibrium. Consequently, current rapid climate changes can disrupt spatially consistent diversity–energy relationships. To understand diversity dynamics, it is crucial to simultaneously consider both the spatial and temporal dimensions of diversity variation.     

Dynamic flux balance analysis of the metabolism of Saccharomyces cerevisiae during the shift from fully respirative or respirofermentative metabolic states to anaerobiosis

Dynamic flux balance analysis was utilized to simulate the metabolic behaviour of initially fully respirative and respirofermentative steady-state cultures of Saccharomyces?cerevisiae during sudden oxygen depletion. The hybrid model for the dynamic flux balance analysis included a stoichiometric genome-scale metabolic model as a static part and dynamic equations for the uptake of glucose and the cessation of respirative metabolism. The yeast consensus genome-scale metabolic model [Herrg?rd MJ et?al. (2008) Nat Biotechnol26, 1155-1160; Dobson PD et?al. (2010) BMC Syst Biol4, 145] was refined with respect to oxygen-dependent energy metabolism and further modified to reflect S.?cerevisiae anabolism in the absence of oxygen. Dynamic flux balance analysis captured well the essential features of the dynamic metabolic behaviour of S.?cerevisiae during adaptation to anaerobiosis. Modelling and simulation enabled the identification of short time-scale flux distribution dynamics under the transition to anaerobic metabolism, during which the specific growth rate was reduced, as well as longer time-scale process dynamics when the specific growth rate recovered. Expression of the metabolic genes was set into the context of the identified dynamics. Metabolic gene expression responses associated with the specific growth rate and with the cessation of respirative metabolism were distinguished.     

Do stresses encountered during the smoked salmon process influence the survival of the spoiling bacterium Shewanella putrefaciens?

The influence of different treatments (i.e. cold, NaCl, phenol and anaerobiosis) encountered during the smoked salmon process was studied by analysing the survival capacity of two Shewanella putrefaciens strains (CIP 69.29 and J13.1). Our results indicated that only the salt stress was critical for the survival of S. putrefaciens. Nevertheless, both strains of S. putrefaciens grown at low temperatures developed a cross-protection to a lethal NaCl treatment. To our knowledge, this is the first report showing that growth at low temperatures induces cross-protection towards NaCl challenge. Moreover, we observed a significant sensitization by moderate salt concentration to a phenol treatment. From our combined data, we propose that control of S. putrefaciens proliferation could take place during the smoked salmon process rather than during storage of the final product.     

A genome-guided analysis of energy conservation in the thermophilic,cytochrome-free acetogenic bacterium Thermoanaerobacter?kivui

Background

Acetogenic bacteria are able to use CO₂ as terminal electron acceptor of an anaerobic respiration, thereby producing acetate with electrons coming from H₂. Due to this feature, acetogens came into focus as platforms to produce biocommodities from waste gases such as H₂ + CO₂ and/or CO. A prerequisite for metabolic engineering is a detailed understanding of the mechanisms of ATP synthesis and electron-transfer reactions to ensure redox homeostasis. Acetogenesis involves the reduction of CO₂ to acetate via soluble enzymes and is coupled to energy conservation by a chemiosmotic mechanism. The membrane-bound module, acting as an ion pump, was of special interest for decades and recently, an Rnf complex was shown to couple electron flow from reduced ferredoxin to NAD⁺ with the export of Na⁺ in Acetobacterium woodii. However, not all acetogens have rnf genes in their genome. In order to gain further insights into energy conservation of non-Rnf-containing, thermophilic acetogens, we sequenced the genome of Thermoanaerobacter kivui.

Results

The genome of Thermoanaerobacter kivui comprises 2.9 Mbp with a G + C content of 35% and 2,378 protein encoding orfs. Neither autotrophic growth nor acetate formation from H₂ + CO₂ was dependent on Na⁺ and acetate formation was inhibited by a protonophore, indicating that H⁺ is used as coupling ion for primary bioenergetics. This is consistent with the finding that the c subunit of the F₁F_O ATP synthase does not have the conserved Na⁺ binding motif. A search for potential H⁺-translocating, membrane-bound protein complexes revealed genes potentially encoding two different proton-reducing, energy-conserving hydrogenases (Ech).

Conclusions

The thermophilic acetogen T. kivui does not use Na⁺ but H⁺ for chemiosmotic ATP synthesis. It does not contain cytochromes and the electrochemical proton gradient is most likely established by an energy-conserving hydrogenase (Ech). Its thermophilic nature and the efficient conversion of H₂ + CO₂ make T. kivui an interesting acetogen to be used for the production of biocommodities in industrial micobiology. Furthermore, our experimental data as well as the increasing number of sequenced genomes of acetogenic bacteria supported the new classification of acetogens into two groups: Rnf- and Ech-containing acetogens.     

Does biomedicine entail the successful reduction of pathology to biology?

The idea of reducing pathology to biology has an extensive history, and the initial forms of the enterprise were unsuccessful. This article discusses the philosophical literature surrounding the notion of reduction in the sciences in general and of biology in particular; reviews several 19th-century programs that promoted the reduction of medicine to other biological disciplines; and examines the post-war origins of the notion of biomedicine. It shows how biology and medicine tend to interact in the constitution of new biomedical knowledge and how the notion of a pathological process resulting in a lesion remains central to the understanding of disease. The article proposes that while strict reduction has yet to be realized, one can speak of a continuing and successful realignment of biology and pathology since the Second World War.     

Who controls the ATP supply in cancer cells? Biochemistry lessons to understand cancer energy metabolism

Applying basic biochemical principles, this review analyzes data that contrasts with the Warburg hypothesis that glycolysis is the exclusive ATP provider in cancer cells. Although disregarded for many years, there is increasing experimental evidence demonstrating that oxidative phosphorylation (OxPhos) makes a significant contribution to ATP supply in many cancer cell types and under a variety of conditions. Substrates oxidized by normal mitochondria such as amino acids and fatty acids are also avidly consumed by cancer cells. In this regard, the proposal that cancer cells metabolize glutamine for anabolic purposes without the need for a functional respiratory chain and OxPhos is analyzed considering thermodynamic and kinetic aspects for the reductive carboxylation of 2-oxoglutarate catalyzed by isocitrate dehydrogenase. In addition, metabolic control analysis (MCA) studies applied to energy metabolism of cancer cells are reevaluated. Regardless of the experimental/environmental conditions and the rate of lactate production, the flux-control of cancer glycolysis is robust in the sense that it involves the same steps: glucose transport, hexokinase, hexosephosphate isomerase and glycogen degradation, all at the beginning of the pathway; these steps together with phosphofructokinase 1 also control glycolysis in normal cells. The respiratory chain complexes exert significantly higher flux-control on OxPhos in cancer cells than in normal cells. Thus, determination of the contribution of each pathway to ATP supply and/or the flux-control distribution of both pathways in cancer cells is necessary in order to identify differences from normal cells which may lead to the design of rational alternative therapies that selectively target cancer energy metabolism.     

A “Thermodynamic” Model of Central Commands Coming to the Muscles during Upper Limb Movements

Neurophysiology - A model of central commands (CCs) coming to the muscles during limb movements is proposed; it is based on experimental studies of the nonlinear dynamics of muscle contraction and...     

Heat sensitivity of eggs attributes to the reduction in Agasicles hygrophila population

Agasicles hygrophila has been introduced worldwide as a control agent for the invasive weed Alternanthera philoxeroides.However,global warming has potential impact on its controlling efficacy.The aim of this research was to explore the primary factors responsible for the greatly reduced A.hygrophila population in hot summers.To imitate the temperature conditions in summers,different developmental stages of hygrophila were treated with high temperatures from 32.5℃ to 45℃ for 1-5 h.Based on the survival rate,the heat tolerance of each developmental stage was ranked from lowest to highest as follows:egg,1st,2nd,3rd instar larva,adult and pupa.Eggs showed the lowest heat tolerance with 37.5℃ as the critical temperature affecting larval hatching.Heat treatment of the A.hygrophila eggs at 37.5℃ for 1 h decreased the hatch rate to 24%.Our results indicated that when compared with the control at 25℃,1 h treatment at 37.5℃ prolonged the duration of the egg stage,shortened the duration of oviposition and total longevity,and changed the reproductive pattern of A.hygrophila.The net reproductive rate,intrinsic rate and finite rate were all significantly reduced.The results suggest that low heat tolerance of the eggs was the major factor responsible for the reduction of A.hygrophila populations,and the key temperature was 37.5°C.Therefore,appropriate measures should be taken to protect eggs in order to maintain the efficacy of A.hygrophila in the biological control of A.philoxeroides in hot summers.