Bacterial ethylene synthesis from 2-oxo-4-thiobutyric acid and from methionine

The ability of selected bacterial cultures to synthesize ethylene during growth in nutrient broth supplemented with methionine or 2-oxo-4-methylthiobutyric acid (KMBA) was examined. Although most cultures transformed KMBA into ethylene, only those of Escherichia coli SPAO and Chromobacterium violaceum were able to convert exogenously added methionine to ethylene. In chemically defined media, E. coli SPAO produced the highest amounts of ethylene from methionine and KMBA. This capability was affected by the nature of the carbon source and the type and amount of nitrogen source used for growth. When glutamate was used as sole source of carbon and nitrogen for growth, the activity of the ethylenogenic enzymes was reduced to 25% of that observed with cultures grown with glucose and NH4Cl. Neither methionine nor KMBA significantly affected the ethylenogenic capacity of E. coli SPAO. Menadione and paraquat, compounds that generate superoxide radicals, stimulated ethylene synthesis by harvested cells, but not by cell-free extracts of E. coli SPAO. In addition, cells of Pseudomonas aeruginosa, which produced no ethylene in culture in the presence of exogenously added KMBA, yet possessed the necessary enzymes in an active form, were able to synthesize ethylene from KMBA when incubated with menadione or paraquat.     

Ethylene production and toxigenicity of methionine and its derivatives with riboflavin in cultures of Verticillium, Fusarium and Colletotrichum species exposed to light

Ethylene was produced by Verticillium dahliae Kleb. grown in liquid Czapek's medium. The rate of ethylene production was enhanced by light but was not affected by shaking or the growth rate of the cultures. L-, D- and DL-methionine, DL-ethionine and a -keto- y -methylthiobutyric acid (KMBA) were good substrates for ethylene production. KMBA may be an intermediate in ethylene production and it appears to be degraded to ethylene either enzymatically by peroxidase or photochemically in the presence of riboflavin. Addition of riboflavin or briefly heating the cultures to 100°C enhanced ethylene production greatly, while the addition of sodium azide, potassium cyanide and catalase were very inhibitory. The SS4 (non-defoliating) pathotype of V. dahliae produced significantly more ethylene (up to 108.4 nl ethylene h₁ from 20 ml-10-day-old cultures) than did the T9 (defoliating) pathotype with all substrates tested. The results suggest that the in vitro rate of ethylene production is not related to the relative virulence of pathotypes of V. dahliae on cotton. A number of Verticillium species, Fusarium oxysporum f. sp. vasinfectum and Colletotrichum dematium var. truncatum were able to produce ethylene in liquid Czapek's medium containing 1 m M L-methionine under continuous light. Riboflavin, although highly stimulatory to ethylene production, caused a fungicidal reaction to all the fungi tested in Czapek's medium containing L-methionine under continuous light. The fungicidal effect of the riboflavin-methionine-light combination occurred at concentrations of riboflavin and methionine less than 1.33 μ M and 0.5 m M , respectively. No fungicidal activity was detected when the cultures were grown in total darkness or when either methionine or riboflavin was omitted from the culture medium.     

Catabolism of volatile sulfur compounds precursors by Brevibacterium linens and Geotrichum candidum, two microorganisms of the cheese ecosystem

Two Brevibacterium linens strains and the cheese-ripening yeast Geotrichum candidum were compared with regard to their ability to produce volatile sulfur compounds (VSCs) from three different precursors namely L-methionine, 4-methylthio-2-oxobutyric acid (KMBA) and 4-methylthio-2-hydroxybutyric acid (HMBA). All microorganisms were able to convert these precursors to VSCs. However, although all were able to produce VSCs from L-methionine, only G. candidum accumulated KMBA when cultivated on this amino acid, contrary to B. linens suggesting that the transamination pathway is not active in this microorganism. Conversely, a L-methionine gamma-lyase activity--which catalyses the one step L-methionine to methanethiol (MTL) degradation route--was only found in B. linens strains. Several other enzymatic activities involved in the catabolism of the precursors tested were investigated. KMBA transiently accumulated in G. candidum cultures, and was then reduced to HMBA by a KMBA dehydrogenase (KDH) activity. This activity was not detected in B. linens. Despite no HMBA dehydrogenase (HDH) was found in G. candidum, a strong HMBA oxidase (HOX) activity was measured in this microorganism. This latter activity was weakly active in B. linens. KMBA and HMBA demethiolating activities were found in all the microorganisms. Our results illustrate the metabolic diversity between cheese-ripening microorganisms of the cheese ecosystem.     

Ethylene formation by cultures of Escherichia coli

Growth of Escherichia coli strain B SPAO on a medium containing glucose, NH₄Cl and methionine resulted in production of ethylene into the culture headspace. When methionine was excluded from the medium there was little formation of ethylene. Ethylene formation in methionine-containing medium occurred for a brief period at the end of exponential growth. Ethylene formation was stimulated by increasing the medium concentration of Fe³⁺ when it was chelated to EDTA. Lowering the medium phosphate concentration also appeared to stimulate ethylene formation. Ethylene formation was inhibited in cultures where NH₄Cl remained in the stationary phase. Synthesis of the ethylene-forming enzyme system was determined by harvesting bacteria at various stages of growth and assaying the capacity of the bacteria to form ethylene from methionine. Ethylene forming capacity was greatest in cultures harvested immediately before and during the period of optimal ethylene formation. It is concluded that ethylene production by E. coli exhibits the typical properties of secondary metabolism.Abbreviations HMBA 2-Hydroxy-4-methylthiobutyric acid (methionine hydroxy analogue) - KMBA 2-keto-4-methylthiobutyric acid - MOPS 3-[N-morpholino] propanesulphonic acid     

Regulation of tyramine oxidase synthesis in Klebsiella aerogenes.

Tyramine oxidase in Klebsiella aerogenes is highly specific for tyramine, dopamine, octopamine, and norepinephrine, and its synthesis is induced specifically by these compounds. The enzyme is present in a membrane-bound form. The Km value for tyramine is 9 X 10(-4) M. Tyramine oxidase synthesis was subjected to catabolite repression by glucose in the presence of ammonium salts. Addition of cyclic adenosine 3',5'-monophosphate (cAMP) overcame the catabolite repression. A mutant strain, K711, which can produce a high level of beta-galactosidase in the presence of glucose and ammonium chloride, can also synthesize tyramine oxidase and histidase in the presence of inducer in glucose ammonium medium. Catabolite repression of tyramine oxidase synthesis was relieved when the cells were grown under conditions of nitrogen limitation, whereas beta-galactosidase was strongly repressed under these conditions. A cAMP-requiring mutant, MK54, synthesized tyramine oxidase rapidly when tyramine was used as the sole source of nitrogen in the absence of cAMP. However, a glutamine synthetase-constitutive mutant, MK94, failed to synthesize tyramine oxidase in the presence of glucose and ammonium chloride, although it synthesized histidase rapidly under these conditions. These results suggest that catabolite repression of tyramine oxidase synthesis in K. aerogenes is regulated by the intracellular level of cAMP and an unknown cytoplasmic factor that acts independently of cAMP and is formed under conditions of nitrogen limitation.     

The effect of indole-3-Acetic acid on ethylene formation in wheat seedlings

Isoperoxidase B 1 isolated from winter wheat (Triticum aestivum L., cv. Jubilar) seedlings was shown to catalyze ethylene formation from α-keto, γ-methylmercaptobutyric acid (KMBA). In the presence of Mn²⁺, indole-3-acetic acid (IAA), andp-coumaric acid, the kinetics by isoperoxidase B 1 catalyzed conversion of KMBA into ethylene and other products was similar to that of IAA oxidation. The reaction rate was therefore controlled by IAA through its electrondonating properties. Exogenous IAA induced ethylene formation in the segments of etiolated wheat coleoptiles. IAA-induced ethylene production was enhanced by L-methionine and mitomycin C. Aminoethoxy-analogue of rhizobitoxine, ferulic acid, sodium benzoate, cycloheximide and actinomyoin D exhibited significant inhibitory effects. These data indicate that the overall reaction mechanism in coleoptile segments involves RNA and protein synthesis. The site of IAA action is not specific; 2,4-dichlorophenoxyacetic, α-naphthylacetic and indole-3-butyric acids, respectively, possessed comparable inductive effect as IAA. Indole-3-propionic acid, indole, L-tryptophan and glucobrassicin had only low inductive efficiency, and moreover indole and L-tryptophan slowed down IAA-induced ethylene formation.     

Glycogen metabolism in resting and growing cells of Saccharomyces carlsbergensis

Saccharomyces carlsbergensis cells, growing under carbohydrate or nitrogen limitation, initially deplete their glycogen, which is resynthesized only during the late exponential phase. Cells, harvested in the carly exponential phase, are even unable to synthesize glycogen in glucose-containing phosphate buffer. This is in contrast to cells from the stationary phase which rapidly synthesize glycogen under the same conditions. Lack of O₂ slows down glycogen synthesis.Contrary to cells suspended in complete medium, addition of ammonia alone to nitrogen free-media induced neither breakdown of glycogen, nor complete cessation of glycogen synthesis. Ammonia slowed down glycogen synthesis (both aerobic and anaerobic), only, in cells grown either under carbohydrate or under nitrogen limitation.Glycogen synthesis was observed 1 min after addition of glucose to a starved cell suspension in phosphate buffer. Removal of the sugar from the buffer resulted in an instantanous decrease of the glycogen level in the cells. The results indicate that glycogen-metabolism is regulated by a variety of endogenous and environmental factors.     

Ethylene formation by cell-free extracts of Escherichia coli

The pathway leading to the formation of ethylene as a secondary metabolite from methionine by Escherichia coli strain B SPAO has been investigated. Methionine was converted to 2-oxo-4-methylthiobutyric acid (KMBA) by a soluble transaminase enzyme. 2-Hydroxy-4-methylthiobutyric acid (HMBA) was also a product, but is probably not an intermediate in the ethylene-forming pathway. KMBA was converted to ethylene, methanethiol and probably carbon dioxide by a soluble enzyme system requiring the presence of NAD(P)H, Fe³⁺ chelated to EDTA, and oxygen. In the absence of added NAD(P)H, ethylene formation by cell-free extracts from KMBA was stimulated by glucose. The transaminase enzyme may allow the amino group to be salvaged from methionine as a source of nitrogen for growth. As in the plant system, ethylene produced by E. coli was derived from the C-3 and C-4 atoms of methionine, but the pathway of formation was different. It seems possible that ethylene production by bacteria might generally occur via the route seen in E. coli.Abbreviations EDTA ethylenediaminetetraacetic acid - HMBA 2-hydroxy-4-methylthiobutyric acid (methionine hydroxy analogue) - HSS high speed supernatant - KMBA 2-oxo-4-methylthiobutyric acid - PCS phase combining system     

The consequence of stimulating glucose dehydrogenase activity by the addition of PQQ on metabolite production by Agrobacterium radiobacter NCIB 11883

Summary Agrobacterium radiobacter NCIB 11 883 does not produce gluconate under conditions of glucose excess in batch or continuous culture. However, the addition of micromolar concentrations of pyrrolo quinoline quinone (PQQ) to fermentation media resulted in rapid excretion of gluconate by batch and continuous cultures. This rapid dehydrogenation of glucose was found in cells grown under carbon and nitrogen limitation and is constitutive which suggests that the only reason why this activity is not normally expressed is due to the inability of the organism to synthesize the prosthetic group (PQQ) of the glucose dehydrogenase enzyme.Although the addition of PQQ to batch and continuous cultures caused a very rapid specific rate of gluconate production (0.6–1.1 g gluconate g^-1 dry wt. h^-1) the rate of exopolysaccharide production remained unaltered. Indeed, when the rates of substrate and oxygen uptake are corrected for the rate of gluconate production in the presence of PQQ there appears to be little physiological consequence as a result of this oxidation.     

Transient response of Enterobacter aerogenes under a dual nutrient limitation in a chemostat.

Utilizing a chemostat with a dual nutrient limitation of nitrogen and phosphate, we examined the transient response of the culture following a pulse of one of the limiting nutrients (ammonia). This method provided quantitative evidence that cells can be grown under dual nutrient limitation. Furthermore, the pattern of response was consistent with the hypothesis that phosphate limitation restricts nucleic acid synthesis in the cell and that nitrogen limitation restricts protein synthesis. The net result is that under a phosphate limitation there is a restricted biosynthetic capacity which we feel is closely associated with the RNA content of the cell.     

Factors affecting poly-β-hydroxybutyrate accumulation in cyanobacteria and in purple non-sulfur bacteria

Abstract Spirulina maxima and Rhodopseudomonas palustris , which are known to be capable of synthesizing poly-β-hydroxybutyrate (PHB), were grown under different conditions in order to investigate the metabolic significance of PHB synthesis in phototrophic microorganisms. The intracellular concentration of PHB in S. maxima , growing photoautotrophically in batch cultures under either balanced or unbalanced (depletion of nitrogen or phosphorus in the mineral medium) conditions, was below 0.005% of cell dry weight. PHB was synthesized (up to 0.7% of dry weight) only after a prolonged period of N-starvation (although no PHB synthesis occurred when N-starvation was induced by azaserine addition) or when cells, after the exhaustion of intracellular phosphorus reserves, became P-starved. Under the latter condition, the PHB concentration reached a value of 1.2% of cell dry weight, the same figure reached in the presence of the uncoupler carbonylcyanide- m -chlorophenylhydrazone (CCCP). When photosynthetic activity was enhanced by a sudden shift of the culture to higher light intensity or when S. maxima was grown at 18°C, no PHB synthesis was detectable. Under all the photoautotrophic growth conditions tested, glycogen was much more heavily accumulated than PHB. Batch cultures of R. palustris , growing photoheterotrophically on acetate with varying nitrogen sources and regimens of nitrogen supplementation, demonstrated that some competition for reducing equivalents exists between nitrogenase activity and PHB biosynthetic pathway. The results seem to suggest that, in phototrophic bacteria able to synthesize both PHB and glycogen, the polyester acts mainly as a regulator of the intracellular reduction charge.     

The effect of pH on nitrogen supply, cell lysis, and solvent production in fermentations of Clostridium acetobutylicum

In batch fermentations of C. acetobutylicum, with 5 g/L yeast extract and 50mM glucose, the ratio of ammonium to glucose affected solvent production when the pH was left to vary uncontrolled from 4.5 to 3.65. High solvent production was observed for a low ratio. When the pH was controlled at 4.5, only acids were produced for all ratio values. At a low ammonium-to-glucose ratio, solvents were produced when the pH was controlled at 3.7. Acids only were produced for a low ratio value at pH 4.0 or for a high ratio value at pH 3.7. In continuous cultures, mostly acids were produced under glucose limitation, but solvents were produced under nitrogen limitation. It was concluded that the nitrogen availability controls solvent production and that the pH affects the availability of organic nitrogen. Biomass autolysis at the stationary phase of batch cultures was reversibly inhibited at pH values less than 3.8. In batch fermentations, the overall molar growth yields on ATP (Y(ATP)) varied from 5.5 to 9.0 and the transient yields from 5.5 to 15.5. In continuous cultures, the Y(ATP) values varied from 5.5 to 14.7 under glucose limitation, and from 6.1 to 9.3 under nitrogen limitation. The Y(ATP) depended on the ammonium to glucose ratio and the culture pH, but did not show the usual dependence on the specific growth rate in batch cultures. The experiments seem to confirm the hypothesis that solvent production is controlled by the demand and availability of ATP.     

Biosynthesis of Poly-beta-Hydroxyalkanoates from Pentoses by Pseudomonas pseudoflava

The potential of Pseudomonas pseudoflava to produce poly-beta-hydroxyalkanoates (PHAs) from pentoses was studied. This organism was able to use a hydrolysate from the hemicellulosic fraction of poplar wood as a carbon and energy source for its growth. However, in batch cultures, growth was inhibited completely at hydrolysate concentrations higher than 30% (vol/vol). When P. pseudoflava was grown on the major sugars present in hemicelluloses in batch cultures, poly-beta-hydroxybutyric acid (PHB) accumulated when glucose, xylose, or arabinose was the sole carbon source, with the final PHB content varying from 17% (wt/wt) of the biomass dry weight on arabinose to 22% (wt/wt) of the biomass dry weight on glucose and xylose. Specific growth rates were 0.58 h on glucose, 0.13 h on xylose, and 0.10 h on arabinose, while the specific PHB production rates based on total biomass ranged from 0.02 g g h on arabinose to 0.11 g g h on glucose. PHB weight-average molecular weights were 640,000 on arabinose and 1,100,000 on glucose and xylose. The absolute amount of PHB in the cells decreased markedly when nitrogen limitation was relaxed by feeding ammonium sulfate at the end of the PHB accumulation stage of the arabinose and xylose fermentations. Copolymers of beta-hydroxybutyric and beta-hydroxyvaleric acids were produced when propionic acid was added to shake flasks containing 10 g of glucose liter. The beta-hydroxyvaleric acid monomer content attained a maximum of 45 mol% when the initial propionic acid concentration was 2 g liter.     

Influence of the nitrogen source on Saccharomyces cerevisiae anaerobic growth and product formation.

To prevent the loss of raw material in ethanol production by anaerobic yeast cultures, glycerol formation has to be reduced. In theory, this may be done by providing the yeast with amino acids, since the de novo cell synthesis of amino acids from glucose and ammonia gives rise to a surplus of NADH, which has to be reoxidized by the formation of glycerol. An industrial strain of Saccharomyces cerevisiae was cultivated in batch cultures with different nitrogen sources, i.e., ammonium salt, glutamic acid, and a mixture of amino acids, with 20 g of glucose per liter as the carbon and energy source. The effects of the nitrogen source on metabolite formation, growth, and cell composition were measured. The glycerol yields obtained with glutamic acid (0.17 mol/mol of glucose) or with the mixture of amino acids (0.10 mol/mol) as a nitrogen source were clearly lower than those for ammonium-grown cultures (0.21 mol/mol). In addition, the ethanol yield increased for growth on both glutamic acid (by 9%) and the mixture of amino acids (by 14%). Glutamic acid has a large influence on the formation of products; the production of, for example, alpha-ketoglutaric acid, succinic acid, and acetic acid, increased compared with their production with the other nitrogen sources. Cultures grown on amino acids have a higher specific growth rate (0.52 h-1) than cultures of both ammonium-grown (0.45 h-1) and glutamic acid-grown (0.33 h-1) cells. Although the product yields differed, similar compositions of the cells were attained. The NADH produced in the amino acid, RNA, and extracellular metabolite syntheses was calculated together with the corresponding glycerol formation. The lower-range values of the theoretically calculated yields of glycerol were in good agreement with the experimental yields, which may indicate that the regulation of metabolism succeeds in the most efficient balancing of the redox potential.     

Effects of growth rate and oxygen tension on glucose dehydrogenase activity inAcinetobacter calcoaceticus LMD 79.41

The regulation of the synthesis of the quinoprotein glucose dehydrogenase (EC 1.1.99.17) has been studied inAcinetobacter calcoaceticus LMD 79.41, an organism able to oxidize glucose to gluconic acid, but unable to grow on both compounds. Glucose dehydrogenase was synthesized constitutively in both batch and carbon-limited chemostat cultures on a variety of substrates. In acetate-limited chemostat cultures glucose dehydrogenase levels and the glucose-oxidizing capacity of whole cells were dependent on the growth rate. They strongly increased at low growth rates at which the maintenance requirement of the cells had a pronounced effect on biomass yield. Cultures grown on a mixture of acetate and glucose in carbon and energy-limited chemostat cultures oxidized glucose quantitatively to gluconic acid. However, during oxygen-limited growth on this mixture glucose was not oxidized and only very low levels of glucose dehydrogenase were detected in cell-free extracts. After introduction of excess oxygen, however, cultures or washed cell suspensions almost instantaneously gained the capacity to oxidize glucose at a high rate, by an as yet unknown mechanism.     

Effect of substrate-dependent microbialy produced ethylene on plant growth

Various compounds have been identified as precursors/substrates for the synthesis of ethylene (C2H4) in soil. This study was designed to compare the efficiency of four substrates, namely L-methionine (L-MET), 2-keto-4-methylthiobutyric acid (KMBA), 1-aminocyclopropane-1-carboxylic acid (ACC), and calcium carbide (CaC2) for ethylene biosynthesis in a sandy clay loam soil by gas chromatography. The classic "triple" response in etiolated pea seedling was employed as a bioassay to demonstrate the effect of substrate-dependent microbialy produced ethylene on plant growth. Results revealed that an amendment with L-MET, KMBA, ACC (up to 0.10 g/kg soil) and CaC2 (0.20 g/kg soil) significantly stimulated ethylene biosynthesis in soil. Overall, ACC proved to be the most effective substrate for ethylene production (1434 nmol/kg soil), followed by KMBA, L-MET, and CaC2 in descending order. Results further revealed that ethylene accumulation in soil released from these substrates created a classic "triple" response in etiolated pea seedlings with different degrees of efficacy. A more obvious classic "triple" response was observed at 0.15, 0.10, and 0.20 g/kg soil of L-MET, KMBA/ACC, and CaC2, respectively. Similarly, direct exposure of etiolated pea seedlings to commercial ethylene gas also modified the growth pattern in the same way. A significant direct correlation (r = 0.86 to 0.97) between substrate-derived [C2H4] and the classic triple response in etiolated pea seedlings was observed. This study demonstrated that the presence of substrate(s) in soil may lead to increased ethylene concentration in the air of the soil, which may affect plant growth in a desired direction.     

Enhancement of chemotaxis in Spirochaeta aurantia grown under conditions of nutrient limitation.

Spirochaeta aurantia M1 cells were grown in a chemostat under conditions of energy and carbon source limitation. The chemotactic responses of the chemostat-grown cells were compared with those of S. aurantia cells grown in batch culture in the presence of excess energy and carbon source. Chemotactic responses were measured by determining the number of cells that entered a capillary tube containing a solution of attractant. S. aurantia cells grown in the chemostat under energy and carbon source limitation exhibited enhanced chemotactic responses and detected lower concentrations of attractant, as compared with cells grown in batch culture. The chemotactic response toward an attractant was specifically enhanced when that attractant was the growth-limiting energy and carbon source. The medium used contained either D-glucose or D-xylose as the sole energy and carbon source. Cells had the greatest chemotactic response toward glucose when grown at a dilution rate (D) of 0.045 h-1 under glucose limitation and toward xylose when grown at D = 0.06 h-1 under xylose limitation. When cells were grown under glucose limitation (D = 0.045 h-1), they sensed concentrations of attractant (glucose) ca. 1,000 times lower than those sensed by batch-grown cells. A similar enhancement of sensing ability (toward xylose) was observed in cells grown under xylose limitation. The results indicated that S. aurantia cells are able to regulate their chemosensory system in response to nutrient limitation. Maximum enhancement of chemotaxis occurs in cells growing at very low concentrations of energy and carbon source. Most likely, this property provides the spirochetes with competitive advantages when the availability of nutrients becomes severely limited in their habitats.     

Biosynthesis and characterization of polyhydroxyalkanoates in the polysaccharide-degrading marine bacterium <Emphasis Type="Italic">Saccharophagus degradans</Emphasis> ATCC 43961

The marine bacterium Saccharophagus degradans was investigated for the synthesis of polyhydroxyalkanoates (PHAs), using glucose as the sole source of carbon in a two-step batch culture. In the first step the microorganism grew under nutrient balanced conditions; in the second step the cells were cultivated under limitation of nitrogen source. The biopolymer accumulated in S. degradans cells was detected by Nile red staining and FT-IR analysis. From GC-MS analysis, it was found that this strain produced a homopolymer of 3-hydroxybutyric acid. The cellular polymer concentration, its molecular mass, glass transition temperature, melting point and heat of fusion were 17.2+/-2.7% of dry cell weight, 54.2+/-0.6 kDa, 37.4+/-6.0 degrees C, 165.6+/-5.5 degrees C and 59.6+/-2.2 J g(-1), respectively. This work is the first report determining the capacity of S. degradans to synthesize PHAs.     

Production and Catabolite Repression of the Constitutive Polygalacturonic Acid trans-Eliminase of Aeromonas liquefaciens

Production of polygalacturonic acid (PGA) trans-eliminase was greatly stimulated under conditions of restricted growth of Aeromonas liquefaciens. This was accomplished either by substrate restriction in a continuous-feeding culture or by restricting divalent cations in a batch culture, with the use of PGA as the sole source of carbon in a chemically defined medium containing inorganic nitrogen. Slow feeding of glucose, glycerol, or PGA to carbon-limited cultures allowed PGA trans-eliminase to be formed at a maximum differential rate 500 times greater than in batch cultures with excess substrate present. The differential rate of enzyme formation obtained by slow feeding of these three substrances or of a mixture of PGA plus glucose was observed to be the same. Therefore, PGA trans-eliminase produced by A. liquefaciens, contrary to the current view, appears to be constitutive. These observations also indicate that production of PGA trans-eliminase is subject to catabolite repression and that limiting the substrate reverses this repression. It was also found that, under conditions of unrestricted growth, any compound which the bacteria can use as a source of carbon and energy repressed constitutive PGA trans-eliminase production. The heritable reversal of catabolite repression of PGA trans-eliminase synthesis was demonstrated by isolation of mutant strain Gc-6 which can readily synthesize the constitutive catabolic enzyme PGA trans-eliminase while growing in the presence of excess substrate.     

Regulation of urease and ammonia assimilatory enzymes in Selenomonas ruminantium.

Urease and glutamine synthetase activities in Selenomonas ruminantium strain D were highest in cells grown in ammonia-limited, linear-growth cultures or when certain compounds other than ammonia served as the nitrogen source and limited the growth rate in batch cultures. Glutamate dehydrogenase activity was highest during glucose (energy)-limited growth or when ammonia was not growth limiting. A positive correlation (R = 0.96) between glutamine synthetase and urease activities was observed for a variety of growth conditions, and both enzyme activities were simultaneously repressed when excess ammonia was added to ammonia-limited, linear-growth cultures. The glutamate analog methionine sulfoximine (MSX), inhibited glutamine synthetase activity in vitro, but glutamate dehydrogenase, glutamate synthase, and urease activities were not affected. The addition of MSX (0.1 to 100 mM) to cultures growing with 20 mM ammonia resulted in growth rate inhibition that was dependent upon the concentration of MSX and was overcome by glutamine addition. Urease activity in MSX-inhibited cultures was increased significantly, suggesting that ammonia was not the direct repressor of urease activity. In ammonia-limited, linear-growth cultures, MSX addition resulted in growth inhibition, a decrease in GS activity, and an increase in urease activity. These results are discussed with respect to the importance of glutamine synthetase and glutamate dehydrogenase for ammonia assimilation under different growth conditions and the relationship of these enzymes to urease.