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1.
Bethany L. Denton Leanne E. Diese Jeffrey L. Firkins Timothy J. Hackmann 《Applied and environmental microbiology》2015,81(5):1832-1838
The aim of this study was to determine if rumen protozoa could form large amounts of reserve carbohydrate compared to the amounts formed by bacteria when competing for glucose in batch cultures. We separated large protozoa and small bacteria from rumen fluid by filtration and centrifugation, recombined equal protein masses of each group into one mixture, and subsequently harvested (reseparated) these groups at intervals after glucose dosing. This method allowed us to monitor reserve carbohydrate accumulation of protozoa and bacteria individually. When mixtures were dosed with a moderate concentration of glucose (4.62 or 5 mM) (n = 2 each), protozoa accumulated large amounts of reserve carbohydrate; 58.7% (standard error of the mean [SEM], 2.2%) glucose carbon was recovered from protozoal reserve carbohydrate at time of peak reserve carbohydrate concentrations. Only 1.7% (SEM, 2.2%) was recovered in bacterial reserve carbohydrate, which was less than that for protozoa (P < 0.001). When provided a high concentration of glucose (20 mM) (n = 4 each), 24.1% (SEM, 2.2%) of glucose carbon was recovered from protozoal reserve carbohydrate, which was still higher (P = 0.001) than the 5.0% (SEM, 2.2%) glucose carbon recovered from bacterial reserve carbohydrate. Our novel competition experiments directly demonstrate that mixed protozoa can sequester sugar away from bacteria by accumulating reserve carbohydrate, giving protozoa a competitive advantage and stabilizing fermentation in the rumen. Similar experiments could be used to investigate the importance of starch sequestration. 相似文献
2.
The energy spilling reactions of bacteria and other organisms 总被引:2,自引:0,他引:2
Russell JB 《Journal of molecular microbiology and biotechnology》2007,13(1-3):1-11
3.
Energy-spilling reactions of Streptococcus bovis and resistance of its membrane to proton conductance. 总被引:10,自引:0,他引:10
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Glucose-excess cultures of Streptococcus bovis consumed glucose faster than the amount that could be explained by growth or maintenance, and nongrowing chloramphenicol-treated cells had a rate of glucose consumption that was 10-fold greater than the maintenance rate. Because N,N-dicyclohexylcarbodiimide, an inhibitor of the membrane-bound F1F0 ATPase, eliminated the nongrowth energy dissipation (energy spilling) without a decrease in ATP and the rate of energy spilling could be increased by the protonophore 3,3',4',5-tetrachlorosalicylanilide, it appeared that a futile cycle of protons through the cell membrane was responsible for most of the energy spilling. When the rate of energy spilling was decreased gradually with iodoacetate, there was only a small decrease in the phosphorylation potential (delta G'p) and the theoretical estimate of H+ per ATP decreased from 4.2 to 3.6. On the bases of this ratio of H+ to ATP and the rate of ATP production, the flux of protons (amperage) across the cell membrane was directly proportional to the rate of energy spilling. Amperage values estimated from delta G'p were, however, nearly twice as great as values which were estimated from the heat production (delta H) of the cells [amperage = (0.38 x wattage)/delta p]. The last comparison indicated that only a fraction of the delta G of ATP hydrolysis was harvested by the F1F0 ATPase to pump protons. Both estimates of amperage indicated that the resistance of the cell membrane to proton conductance was inversely proportional to the log of the energy-spilling rate. 相似文献
4.
Jianming Liu Vijayalakshmi Kandasamy Anders Würtz Peter Ruhdal Jensen Christian Solem 《Applied microbiology and biotechnology》2016,100(22):9509-9517
Having a sufficient supply of energy, usually in the form of ATP, is essential for all living organisms. In this study, however, we demonstrate that it can be beneficial to reduce ATP availability when the objective is microbial production. By introducing the ATP hydrolyzing F1-ATPase into a Lactococcus lactis strain engineered into producing acetoin, we show that production titer and yield both can be increased. At high F1-ATPase expression level, the acetoin production yield could be increased by 10 %; however, because of the negative effect that the F1-ATPase had on biomass yield and growth, this increase was at the cost of volumetric productivity. By lowering the expression level of the F1-ATPase, both the volumetric productivity and the final yield could be increased by 5 % compared to the reference strain not overexpressing the F1-ATPase, and in batch fermentation, it was possible to convert 176 mM (32 g/L) of glucose into 146.5 mM (12.9 g/L) acetoin with a yield of 83 % of the theoretical maximum. To further demonstrate the potential of the cell factory developed, we complemented it with the lactose plasmid pLP712, which allowed for growth and acetoin production from a dairy waste stream, deproteinized whey. Using this cheap and renewable feedstock, efficient acetoin production with a titer of 157 mM (14 g/L) acetoin was accomplished. 相似文献
5.
《Animal : an international journal of animal bioscience》2015,9(11):1795-1806
Many feeding trials have been conducted to quantify enteric methane (CH4) production in ruminants. Although a relationship between diet composition, rumen fermentation and CH4 production is generally accepted, the efforts to quantify this relationship within the same experiment remain scarce. In the present study, a data set was compiled from the results of three intensive respiration chamber trials with lactating rumen and intestinal fistulated Holstein cows, including measurements of rumen and intestinal digestion, rumen fermentation parameters and CH4 production. Two approaches were used to calculate CH4 from observations: (1) a rumen organic matter (OM) balance was derived from OM intake and duodenal organic matter flow (DOM) distinguishing various nutrients and (2) a rumen carbon balance was derived from carbon intake and duodenal carbon flow (DCARB). Duodenal flow was corrected for endogenous matter, and contribution of fermentation in the large intestine was accounted for. Hydrogen (H2) arising from fermentation was calculated using the fermentation pattern measured in rumen fluid. CH4 was calculated from H2 production corrected for H2 use with biohydrogenation of fatty acids. The DOM model overestimated CH4/kg dry matter intake (DMI) by 6.1% (R2=0.36) and the DCARB model underestimated CH4/kg DMI by 0.4% (R2=0.43). A stepwise regression of the difference between measured and calculated daily CH4 production was conducted to examine explanations for the deviance. Dietary carbohydrate composition and rumen carbohydrate digestion were the main sources of inaccuracies for both models. Furthermore, differences were related to rumen ammonia concentration with the DOM model and to rumen pH and dietary fat with the DCARB model. Adding these parameters to the models and performing a multiple regression against observed daily CH4 production resulted in R2 of 0.66 and 0.72 for DOM and DCARB models, respectively. The diurnal pattern of CH4 production followed that of rumen volatile fatty acid (VFA) concentration and the CH4 to CO2 production ratio, but was inverse to rumen pH and the rumen hydrogen balance calculated from 4×(acetate+butyrate)/2×(propionate+valerate). In conclusion, the amount of feed fermented was the most important factor determining variations in CH4 production between animals, diets and during the day. Interactions between feed components, VFA absorption rates and variation between animals seemed to be factors that were complicating the accurate prediction of CH4. Using a ruminal carbon balance appeared to predict CH4 production just as well as calculations based on rumen digestion of individual nutrients. 相似文献
6.
Relationship between Intracellular Phosphate, Proton Motive Force, and Rate of Nongrowth Energy Dissipation (Energy Spilling) in Streptococcus bovis JB1
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When the rate of glucose addition to nongrowing Streptococcus bovis cell suspensions was increased, the fermentation was homolactic, fructose-1,6-diphosphate (FDP) increased, intracellular inorganic phosphate (Pi) declined, and the energy-spilling rate increased. ATP and ADP were not significantly affected by glucose consumption rate, but the decrease in Pi was sufficient to cause an increase in the free energy of ATP hydrolysis (ΔG′p). The increase in ΔG′p was correlated with an increase in proton motive force (Δp). S. bovis continuous cultures (dilution rate of 0.65 h−1) that were provided with ammonia as the sole nitrogen source also had high rates of lactate production and energy spilling. When Trypticase was added as a source of amino acids, lactate production decreased; a greater fraction of the glucose was converted to acetate, formate, and ethanol; and the energy-spilling rate decreased. Trypticase also caused a decrease in FDP, an increase in Pi, and a decrease in Δp. The change in Δp could be explained by Pi-dependent changes in the ΔG′p. When Pi declined, ΔG′p and Δp increased. The ratio of ΔG′p to Δp (millivolt per millivolt) was always high (>4) at low rates of energy spilling but declined when the energy-spilling rate increased. Based on these results, it appears that Δp and the energy-spilling rate are responsive to fluctuations in the intracellular Pi concentration. 相似文献
7.
A Role for Fructose 1,6-Diphosphate in the ATPase-Mediated Energy-Spilling Reaction of Streptococcus bovis 总被引:2,自引:1,他引:1
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The amount of ATP produced by Streptococcus bovis was larger than the amount that could be attributed to growth and maintenance, and even glucose-limited continuous cultures used ATP inefficiently (spilled ATP). Rapid-dilution-rate cultures always spilled more ATP than those growing at slow dilution rates, but rates of ATP spilling could also be enhanced by amino acid deprivation (with only ammonia as a nitrogen source). Energy spilling and intracellular ATP were not correlated, but energy spilling was always greatest when the rate of lactate production was high. The relationship between lactate production and energy spilling was supported by the observation that amino acid deprivation increased lactate production and ATP spilling. The lactate production rate of nongrowing (energy-spilling) S. bovis cells was fructose 1,6-diphosphate (FDP) dependent, and previous work showed that the lactate dehydrogenase of S. bovis was activated by FDP (M. J. Wolin, Science 146:775-777, 1964). The role of FDP in energy spilling was supported by the observation that the membrane-bound ATPase of S. bovis could be stimulated by FDP. FDP decreased the K(infm) for ATP by as much as fivefold. Other glycolytic intermediates could not stimulate the ATPase of washed membrane preparations, and FDP had no effect on soluble ATPase activity. 相似文献
8.
The antibody-secreting murine hybridoma, CC9C10, was grown in batch culture in a medium containing 20 mM glucose and 2 mM glutamine. After 2 days of exponential growth, the glutamine content of the medium was completely depleted, whereas the glucose content was reduced to 60% of the original concentration. The glucose and glutamine metabolism was analyzed at midexponential phase by use of radioactively labelled substrates. Glycolysis accounted for the metabolism of most of the glucose utilized (> 96%) with flux through the pentose phosphate pathway (3.6%) and the TCA cycle (0.6%) accounting for the remainder. Glutamine was partially oxidised via glutaminolysis to alanine (55%), aspartate (3%), glutamate (4%), lactate (9%), and CO2 (22%). Calculation of the theoretical ATP production from these pathways indicated that glucose could provide 59% and glutamine 41% of the energy requirement of the cells. © 1994 Wiley-Liss, Inc. 相似文献
9.
In vitro rumen fermentation of soluble and non-soluble polymeric carbohydrates in relation to ruminal acidosis 总被引:1,自引:0,他引:1
The end-products of dietary carbohydrate fermentation catalysed by rumen microflora can serve as the primary source of energy for ruminants. However, ruminants provided with continuous carbohydrate-containing feed can develop a metabolic disorder called “acidosis”. We have evaluated the fermentation pattern of both soluble monomeric and non-soluble polymeric carbohydrates in the rumen in in vitro fermentation trials. We found that acidosis could occur within 6 h of incubation in the rumen culture fermenting sugars and starch. The formation of lactic acid and acetic acid, either alone or in mixture with ethanol, accounted for high build-up of acid in the rumen. Acidosis resulted even when only 20% of a normal daily feed load for all soluble and non-soluble carbohydrates was provided. DNA-based microbial analysis revealed that Prevotella was the dominant microbial species present in the rumen fluid. 相似文献
10.
Summary The heat evolution of aerobic batch cultures of growing yeast (Saccharomyces cerevisiae) in glucose media was investigated by a combination of a flow-microcalorimeter with a fermentor vessel. The course of heat production, cell production and the rate of oxygen consumption were qualitatively the same for all glucose concentrations between 10 mM and 100 mM. Under optimal aerobic conditions a triphasic growth was observed due to the fermentation of glucose to ethanol, respiration of ethanol to CO2 and acetate, and respiration of acetate to C02. Energy and carbon were found to be in balance for all glucose concentrations. 相似文献
11.
P.M. Matthews J.L. Bland D.G. Gadian G.K. Radda 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》1982,721(3):312-320
(1) 31P nuclear magnetic resonance was used to measure the creatine kinase-catalysed fluxes in Langendorff-perfused rat hearts consuming oxygen at different rates and using either of two exogenous substrates (11 mM glucose or 5 mM acetate). (2) Fluxes in the direction of ATP synthesis were between 3.5–12-times the steady-state rates of ATP utilization (estimated from rates of O2-consumption), demonstrating that the reaction is sufficiently rapid to maintain the cytosolic reactants near their equilibrium concentrations. (3) Under all conditions studied, the cytosolic free [ADP] was primarily responsible for regulating the creatine kinase fluxes. The enzyme displayed a Km for cytosolic ADP of 35 μM and an apparent Vmax of 5.5 mM/s in the intact tissue. (4) Although the reaction is maintained in an overall steady-state, the measured ratio of the forward flux (ATP synthesis) to the reverse flux (phosphocreatine synthesis) was significantly greater than unity under some conditions. It is proposed that this discrepancy may be a consequence of participation of ATP in reactions other than the PCr /ag ATP or ATP /ag ADP + Pi interconversions specifically considered in the analysis. (5) The results support the view that creatine kinase functions primarily to maintain low cytosolic concentrations of ADP during transient periods in which energy utilization exceeds production. 相似文献
12.
Bingkun Yan Qingbao Ding Ling Ou Zhi Zou 《World journal of microbiology & biotechnology》2014,30(3):1123-1128
A process of glucose-6-phosphate (G-6-P) production coupled with an adenosine triphosphate (ATP) regeneration system was constructed that utilized acetyl phosphate (ACP) via acetate kinase (ACKase). The genes glk and ack from Escherichia coli K12 were amplified and cloned into pET-28a(+), then transformed into E. coli BL21 (DE3) and the recombinant strains were named pGLK and pACK respectively. Glucokinase (glkase) in pGLK and ACKase in pACK were both overexpressed in soluble form. G-6-P was efficiently produced from glucose and ACP using a very small amount of ATP. The conversion yield was greater than 97 % when the reaction solution containing 10 mM glucose, 20 mM ACP-Na2, 0.5 mM ATP, 5 mM Mg2+, 50 mM potassium phosphate buffer (pH 7.0), 4.856 U glkase and 3.632 U ACKase were put into 37 °C water bath for 1 h. 相似文献
13.
When the ruminal bacterium Streptococcus bovis was grown in batch culture with glucose as the energy source, the doubling time was approximately 21 min and the rate of bacterial heat production was proportional to the optical density (1.72 W/g protein). If exponentially growing cultures were treated with chloramphenicol, there was a decline in heat production, but the rate was greater than 0.30 W/g protein even after growth ceased. Since there was no heat production after glucose depletion, this growth-independent energy dissipation (spilling) was not simply due to endogenous metabolism. Stationary cells which were washed and incubated in nitrogen-free medium containing an excess of glucose produced heat at a rate of 0.17 W/g protein. Monensin and tetrachlorosalicylanilide (TCS), compounds which facilitate an influx of protons, caused a more than 2-fold increase in heat production. Dicyclohexylcarbodiimide (DCCD) virtually eliminated growth-independent heat production regardless of the mode of growth inhibition. Because DCCD had little effect on the glucose phosphotransferase system, it appeared that the combined action of proton influx and the membrane bound F1F0 proton ATPase was responsible for energy spilling.Abbreviations DCCD
dicyclohexylcarbodiimide
- TCS
tetrachlorosalicylanilide 相似文献
14.
Osório H Moradas-Ferreira P Günther Sillero MA Sillero A 《Archives of microbiology》2004,181(3):231-236
As has been previously shown, Saccharomyces cerevisiae grown in 2% or 0.025% glucose uses this carbohydrate by the fermentative or oxidative pathways, respectively. Depending on the glucose concentration in the medium, the effect of the addition of H2O2 on the level of ATP and on glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activity differed. In the presence of 2% glucose, ATP and GAPDH decreased sharply during the first few minutes of treatment, whereas in the presence of 0.025% glucose, GAPDH activity decreased similarly, but the ATP level remained practically unchanged. The addition of 3 mM glutathione to the culture media prevented the depletion of ATP levels and GAPDH activity in the presence of H2O2. Catalase and superoxide dismutase activities did not vary significantly when yeast cells were grown either in 2% or in 0.025% glucose. 相似文献
15.
Use of Adenosine 5′-Triphosphate as an Indicator of the Microbiota Biomass in Rumen Contents
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A number of techniques were tested for their efficiency in extracting adenosine 5′-triphosphate (ATP) from strained rumen fluid (SRF). Extraction with 0.6 N H2SO4, using a modification of the procedure described by Lee et al. (1971), was the most efficient and was better suited for extracting particulate samples. Neutralized extracts could not be stored frozen before assaying for ATP because large losses were incurred. The inclusion of internal standards was necessary to correct for incomplete recovery of ATP. The ATP concentration in rumen contents from a cow receiving a ration of dried roughage (mainly alfalfa hay) ranged from 31 to 56 μg of ATP per g of contents. Approximately 75% of the ATP was associated with the particulate material. The ATP was primarily of microbial origin, since only traces of ATP were present in the feed and none was found in “cell-free” rumen fluid. Fractionation of the bacterial and protozoal populations in SRF resulted in the isolation of an enriched protozoal fraction with a 10-fold higher ATP concentration than that of the separated rumen bacteria. The ATP pool sizes of nine functionally important rumen bacteria during the exponential phase of growth ranged from 1.1 to 17.6 μg of ATP per mg of dry weight. This information indicates that using ATP as a measure of microbial biomass in rumen contents must be done with caution because of possible variations in the efficiency of extraction of ATP from rumen contents and differences in the concentration of ATP in rumen microbes. 相似文献
16.
《Journal of Fermentation Technology》1987,65(3):255-260
Improvement of efficiency of ATP production from adenosine with sorbitol-treated cells of Candida boidinii (Kloeckera sp.) no. 2201 was investigatedOrthophosphate, pyrophosphate and divalent metal ions inhibited the deamination of AMP to IMP, a major by-product. No hypoxanthine nucleotidic compound was accumulated by addition of coformycin. By successive feeding of K2HPO4 to maintain the phosphorus concentration at over 100 mM, the conversion rate from adenosine to ATP was improved to 70%. Simultaneous feeding of K2HPO4 and adenosine resulted in the accumulation of 100 mM ATP (50.7 g/l) after 28 h of incubation and the increase of IMP without decrease of ATP for 48 h.ATP production was further prolonged until 60 h of incubation and 198 mM ATP (100 g/l) was accumulated at the conversion rate of 77.4% by control of pH in the range of 6.5 to 6.8 during the reaction. 相似文献
17.
l-arginine, a semi essential amino acid, is an important amino acid in food flavoring and pharmaceutical industries. Its production by microbial fermentation is gaining more and more attention. In previous work, we obtained a new l-arginine producing Corynebacterium crenatum (subspecies of Corynebacterium glutamicum) through mutation breeding. In this work, we enhanced l-arginine production through improvement of the intracellular environment. First, two NAD(P)H-dependent H2O2-forming flavin reductases Frd181 (encoded by frd1 gene) and Frd188 (encoded by frd2) in C. glutamicum were identified for the first time. Next, the roles of Frd181 and Frd188 in C. glutamicum were studied by overexpression and deletion of the encoding genes, and the results showed that the inactivation of Frd181 and Frd188 was beneficial for cell growth and l-arginine production, owing to the decreased H2O2 synthesis and intracellular reactive oxygen species (ROS) level, and increased intracellular NADH and ATP levels. Then, the ATP level was further increased by deletion of noxA (encoding NADH oxidase) and amn (encoding AMP nucleosidase), and overexpression of pgk (encoding 3-phosphoglycerate kinase) and pyk (encoding pyruvate kinase), and the l-arginine production and yield from glucose were significantly increased. In fed-batch fermentation, the l-arginine production and yield from glucose of the final strain reached 57.3 g/L and 0.326 g/g, respectively, which were 49.2% and 34.2% higher than those of the parent strain, respectively. ROS and ATP are important elements of the intracellular environment, and l-arginine biosynthesis requires a large amount of ATP. For the first time, we enhanced l-arginine production and yield from glucose through reducing the H2O2 synthesis and increasing the ATP supply. 相似文献
18.
Vikas Kumar Patel Deepamala Maji Anil Kumar Singh M. R. Suseela Shanthy Sundaram Alok Kalra 《Journal of applied phycology》2014,26(1):279-286
Cyanobacteria have evolved photosynthetic mechanisms in which solar energy is used to fix CO2 into carbohydrates. The lipids from cyanobacteria can be converted to biodiesel by extraction–transesterification methods. The present study demonstrates the usefulness of the natural plant and microbial growth promoter calliterpenone from the plant Callicarpa macrophylla supplemented at three different doses (15, 25, 50 μL of a 0.01 mM solution) per 100 mL BG11+ medium for enhancing total biomass, carbohydrate, and lipid yields and reducing the surface-to-volume ratios of cells of Synechocystis PCC 6803. The enhanced total dried biomass, carbohydrate, and lipid production was 316.1, 140.34, and 130.76 %, respectively, higher than the control, and were obtained after 15 days of cultivation at the dose of 15 μL (0.01 mM) of calliterpenone per 100 mL BG11+ medium. A decrease in surface-to-volume ratio of cells from 1.19 to 0.84 compared to the control was also observed. Response surface methodology was used to optimize the doses of calliterpenone at different pH of growth media. An increase of 346.95, 187.2, and 134.46 % in biomass, carbohydrate, and lipid yields, respectively, was achieved after 10 days of cultivation in optimized BG11+ media at pH 7.5 and with 20 μL (0.01 mM) calliterpenone per 100 mL. Thus, this biomolecule can be exploited for higher yields of Synechocystis PCC 6803 in a relatively shorter culture time making this an attractive strategy for fuel production using this cyanobacterium. 相似文献
19.
《Biochemical education》1998,26(1):22-23
The authors of many recent popular textbooks of biochemistry quote values for the ‘energy content’ of triacylglcyerol and dry carbohydrate on a weight basis as well as presenting calculations for the yield of ATP obtained when a molecule of glucose, or palmitic acid, is completely oxidised to CO2 and H2O. By extending these calculations and computing the yield of ATP in terms of the weight of glucose or palmitic acid oxidised to CO2 and H2O, it can be shown that the value for the ratio of the energy content of fat to that of carbohydrate is almost identical to the ratio of the yield of ATP per gram of palmitic acid oxidised, compared with that of glucose. Therefore, the efficiency (on a per gram basis) by which energy is made available as ATP is comparable for both the oxidation of fat and carbohydrate, thus underscoring the fact that the catabolic pathways for both fat and carbohydrate ultimately use the same means of generating ATP, namely, oxidative phosphorylation. 相似文献
20.
Caldicellulosiruptor saccharolyticus displays superior H2 yields on a wide range of carbon sources provided that lactate formation is avoided. Nevertheless, a low lactate flux is initiated as the growth rate declined in the transition to the stationary phase, which coincides with a drastic decrease in the glucose consumption and acetate production fluxes. In addition, the decrease in growth rate was accompanied by a sudden increase and then decrease in NADH levels. The V′MAX of the lactate dehydrogenase (LDH) doubled when the cells entered the stationary phase. Kinetic analysis revealed that at the metabolic level LDH activity is regulated through (i) competitive inhibition by pyrophosphate (PPi, ki=1.7 mM) and NAD (ki=0.43 mM) and (ii) allosteric activation by FBP (300%), ATP (160%) and ADP (140%). From these data a MWC-based model was derived. Simulations with this model could explain the observed lactate shift by displaying how the sensitivity of LDH activity to NADH/NAD ratio varied with different PPi concentrations. Moreover, the activation of LDH by ATP indicates that C. saccharolyticus uses LDH as a means to adjusts its flux of ATP and NADH production. To our knowledge, this is the first time PPi is observed as an effector of LDH. 相似文献