Studies on phosphate metabolism in obligate methanotrophs

Trotsenko, Yu. A.; Shishkina, Valentina N.

doi:10.1111/j.1574-6968.1990.tb04923.x

Cited by 33 publications

(12 citation statements)

References 14 publications

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“…Thus, a number of phosphorylated compounds are produced by H. volcanii at the inception of the late exponential phase of growth or after prolonged nutrient or energy starvation. One hypothesis for this phenomenon is that the molecules produced are short-chain polyphosphates, which have been shown to be produced by other microbial organisms (13,18,20). To verify this hypothesis, formic acid extracts were prepared from S. cerevisiae, an organism with well-characterized polyphosphate expression (13,18), and the spots obtained were compared with those produced by H. volcanii.…”

Section: Resultsmentioning

confidence: 99%

Lack of production of (p)ppGpp in Halobacterium volcanii under conditions that are effective in the eubacteria

1995

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The stringent halobacterial strain Haloferax volcanii was subjected to a set of physiological conditions different from amino acid starvation that are known to cause production of guanosine polyphosphates [(p)pp Gpp] in eubacteria via the relA-independent (spoT) pathway. The conditions used were temperature upshift, treatment with cyanide, and total starvation. Under none of these conditions were detectable levels of (p)ppGpp observed. This result, in conjunction with our previous finding that (p)ppGpp synthesis does not occur under amino acid starvation, leads to the conclusion that in halobacteria both growth rate control and stringency are probably governed by mechanisms that operate in the absence of ppGpp. During exponential growth, a low level of phosphorylated compounds with electrophoretic mobilities similar, but not identical, to that of (p)ppGpp were observed. The intracellular concentration of these compounds increased considerably during the stationary phase of growth and with all of the treatments used. The compounds were identified as short-chain polyphosphates identical to those found under similar conditions in Saccharomyces cerevisiae.In 1969, Cashel and Gallant reported that wild-type Escherichia coli is capable of producing two unusual compounds originally named magic spots I and II and later identified as guanosine tetra-and pentaphosphates, respectively [collectively referred to as (p)ppGpp] (3). These guanosine nucleotides are generated in a relA-dependent manner by the relA gene product, (p)ppGpp synthetase I, and in a relA-independent manner by the spoT gene product, the SpoT enzyme, which is also implicated in (p)ppGpp degradation. Kinetic studies have shown that pppGpp is formed from GTP, that ppGpp is derived from pppGpp, and that pppGpp inhibits its own synthesis (12).In eubacteria, the rate of stable RNA synthesis at different growth rates is subject to a control mechanism that is distinct from that of other cellular molecules (growth rate control of RNA synthesis). An inverse correlation between the ppGpp concentration, produced in this case by the relA-independent pathway, and the rate of stable RNA synthesis was observed at a wide range of different growth rates (5), which suggested that ppGpp could act as an effector molecule negatively regulating stable RNA synthesis in growth rate control. The isolation by Xiao et al. (21) of relA spoT mutants unable to produce ppGpp permitted Gaal and Gourse (8) to show that ppGpp either is not the mediator or does not act alone, since growth rate control is normal in the double mutants. Hernandez and Bremer have recently shown that in wild-type E. coli, three factors increase with the growth rate: RNA polymerase concentration, RNA polymerase activity, and the distribution of active RNA polymerase between stable and mRNA genes. In an E. coli⌬relA⌬spoT mutant that does not produce ppGpp, RNA polymerase synthesis and activity increase with the growth rate but the distribution of active RNA polymerase between stable and mRNA genes does not. This...

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Section: Resultsmentioning

confidence: 99%

Lack of production of (p)ppGpp in Halobacterium volcanii under conditions that are effective in the eubacteria

1995

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“…Despite a proposed high phosphate demand (2^25 mM) of Methylosinus trichosporium for growth [34], even concentrations as low as 0.015 mM phosphate proved to be su⁄cient in our study. This contradiction might be explained by the ¢nd-ing that MOB can rely on intracellular polyphosphates [35], which have been accumulated when the MOB were growing in the sediment.…”

Section: E¡ect Of Medium Constituentsmentioning

confidence: 99%

Preferential cultivation of type II methanotrophic bacteria from littoral sediments (Lake Constance)

Bussmann

Pester

Brune

et al. 2004

FEMS Microbiology Ecology

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Most widely used medium for cultivation of methanotrophic bacteria from various environments is that proposed in 1970 by Whittenbury. In order to adapt and optimize medium for culturing of methanotrophs from freshwater sediment, media with varying concentrations of substrates, phosphate, nitrate, and other mineral salts were used to enumerate methanotrophs by the most probable number method. High concentrations ( s 1 mM) of magnesium and sulfate, and high concentrations of nitrate ( s 500 WM) significantly reduced the number of cultured methanotrophs, whereas phosphate in the range of 15^1500 WM had no influence. Also oxygen and carbon dioxide influenced the culturing efficiency, with an optimal mixing ratio of 17% O 2 and 3% CO 2 ; the mixing ratio of methane (63 2%) had no effect. A clone library of pmoA genes amplified by PCR from DNA extracted from sediment revealed the presence of both type I and type II methanotrophs. Nonetheless, the cultivation of methanotrophs, also with the improved medium, clearly favored growth of type II methanotrophs of the Methylosinus/Methylocystis group. Although significantly more methanotrophs could be cultured with the modified medium, their diversity did not mirror the diversity of methanotrophs in the sediment sample detected by molecular biology method.

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“…In general this type of PFK enzyme is associated with anaerobic metabolism (31,32) in Propionibacterium spp. (33), various protozoa (32) and amoebas (35), and higher plants (6 (2,38). The use of PP1 as a phosphate donor thus raises questions about the metabolism of PP1 and its further effects on the regulation of glycolysis inA.…”

Section: Ppi-pfk (Not Shown)mentioning

confidence: 99%

Enzymes of glucose and methanol metabolism in the actinomycete Amycolatopsis methanolica

et al. 1994

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The actinomycete Amycolatopsis methanolica was found to employ the normal bacterial set of glycolytic and pentose phosphate pathway enzymes, except for the presence of a PPi-dependent phosphofructokinase (PPi-PFK) and a 3-phosphoglycerate mutase that is stimulated by 2,3-bisphosphoglycerate. Screening of a number of actinomycetes revealed PP,-PFK activity only in members of the family Pseudonocardiaceae. The A. methanolica PP,-PFK and 3-phosphoglycerate mutase enzymes were purified to homogeneity. PP,-PFK appeared to be insensitive to the typical effectors of ATP-dependent PFK enzymes. Nevertheless, strong N-terminal amino acid sequence homology was found with ATP-PFK enzymes from other bacteria. The A. methanolica pyruvate kinase was purified over 250-fold and characterized as an allosteric enzyme, sensitive to inhibition by P, and ATP but stimulated by AMP. By using mutants, evidence was obtained for the presence of transketolase isoenzymes functioning in the pentose phosphate pathway and ribulose monophosphate cycle during growth on glucose and methanol, respectively.Actinomycetes are important bacterial producers of secondary metabolites. There is a strong interest in the genetics of secondary-metabolite biosynthesis, with most studies concentrating on these pathways and their control. Many secondary metabolites are initially derived from intermediates of the central pathways of primary metabolism. Little is currently known, however, about the enzymes and regulation of, for instance, glucose metabolism in actinomycetes. This is mostly because of a general lack of physiological studies on primary metabolism in actinomycetes (21). We have initiated such studies with the actinomycete Amycolatopsis methanolica (8), belonging to the family Pseudonocardiaceae (42), which includes many species producing bioactive compounds, e.g., the antibiotics rifamycin and erythromycin. A. methanolica is one of the few methanol-utilizing gram-positive bacteria known (10,12,17). Methanol oxidation via formaldehyde and formate to carbon dioxide results in energy generation (5,17). Carbon assimilation starts by formaldehyde fixation via the ribulose monophosphate (RuMP) cycle (9,17). This cycle involves the specific enzymes hexulose-6-phosphate synthase (HPS) and hexulose-6-phosphate isomerase (HPI), the glycolytic enzymes 6-phosphofructokinase (PFK) and fructose-1,6-bisphosphate (FBP) aldolase (9), and various enzymes also involved in the related pentose phosphate pathway (Fig. 1) (12).The identity, properties, and regulation of enzymes involved in glucose and methanol metabolism in A. methanolica were examined in this study. MATERUILS AND METHODSMicroorganisms and cultivation. Wild-type A. methanolica NCIB 11946, its maintenance, and the procedures followed for cultivation in batch cultures, harvesting of cells, and growth measurements have been described previously (8)(9)(10)(11)

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Studies on phosphate metabolism in obligate methanotrophs

Cited by 33 publications

References 14 publications

Lack of production of (p)ppGpp in Halobacterium volcanii under conditions that are effective in the eubacteria

Lack of production of (p)ppGpp in Halobacterium volcanii under conditions that are effective in the eubacteria

Preferential cultivation of type II methanotrophic bacteria from littoral sediments (Lake Constance)

Enzymes of glucose and methanol metabolism in the actinomycete Amycolatopsis methanolica

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