The control of synthesis of bacterial cell walls. Interaction in the synthesis of nucleotide precursors

Anderson, Raymond G.; Douglas, L. Julia; Hussey, Helen; Baddiley, J.

doi:10.1042/bj1360871

Cited by 26 publications

(13 citation statements)

References 19 publications

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“…The results did not allow identification of the factor(s) responsible for the decrease in activity. Nevertheless, comparison with previous results revealed that the lowest figure obtained here of 33 nmol min-I (mg protein)-', measured in the presence of 10-fold higher concentrations of a-groP and CTP than those in the standard assay, is close to that reported (Anderson et al, 1973) for a partially purified preparation of gro-PCT. When assayed under the standard (non-saturating) conditions employed here, the final figure of 7 units is about one-half the value (between 14 and 19 units) reported by Glaser & Loewy (1979) for a partially purified enzyme preparation.…”

Section: Preparation Of'a Crude Cytoso1fi-actionfr Assay Of'gro-pctmentioning

confidence: 63%

A conditional-lethal mutant of Bacillus subtilis 168 with a thermosensitive glycerol-3-phosphate cytidylyltransferase, an enzyme specific for the synthesis of the major wall teichoic acid

Pooley¹,

Abellan²,

Karamata³

1991

Journal of General Microbiology

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A biochemical analysis was undertaken of thermosensitive mutants of BaciZZus subtiZis 168 harbouring mutations in several tag genes, involved in the synthesis of the major wall teichoic acid, poly(glycero1 phosphate), poly(groP). Incorporation of a pulse of [2-3H]glycerol into whole cells, following shift to the restrictive growth temperature, was used to assess synthesis of this polymer and to seek evidence of accumulation of a specific precursor. The rate of incorporation into poly(groP) was strongly decreased in all mutants; glycerol uptake was diminished by 80% or more for a strain harbouring mutation tugs1 (formerly fag-1) and one bearing tagDll (formerly tag-11). The pool of CDP-glycerol (CDP-gro), a specific precursor of poly(groP), was increased, relative to the wild-type, for all mutations except tagDll, where the pool of CDP-gro was reduced. Cytoplasmic extracts, assayed at the permissive temperature for glycerol-3-phosphate cytidylyltransferase (gro-PCT), the enzyme synthesizing CDPgro, revealed wild-type activities for all mutations except tagDl1. Gro-PCT activity in the latter strain was 100-fold lower and, unlike that in all other mutant strains, highly thermolabile. This thermosensitivity suggests that tagD encodes gro-PCT. The identification, in a gene encoding a poly(groP)-specific enzyme, of a mutation conferring a thermosensitive growth phenotype renders explicit the conclusion that synthesis of this teichoic acid is essential for the growth of B. subtiZis.

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Section: Preparation Of'a Crude Cytoso1fi-actionfr Assay Of'gro-pctmentioning

confidence: 63%

A conditional-lethal mutant of Bacillus subtilis 168 with a thermosensitive glycerol-3-phosphate cytidylyltransferase, an enzyme specific for the synthesis of the major wall teichoic acid

Pooley¹,

Abellan²,

Karamata³

1991

Journal of General Microbiology

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“…The standard assay mixture contained 50 mM Tris-hydrochloride buffer (pH 8.6), 1 Purification of GlmU from overproducing strain JM83 (pMLD78). The total extract (67 mg of protein, 552 U of uridyltransferase activity) secured from strain JM83 (pMLD78) as described above was loaded onto a column (12.5 by 2.5 cm) of DEAE-Trisacryl-M (IBF, Vileneuve-laGarenne, France) that had been preequilibrated with buffer A (20 mM potassium phosphate buffer, pH 7.4, containing 1 mM ,B-mercaptoethanol and 10% [vol/vol) glycerol).…”

Section: Methodsmentioning

confidence: 99%

“…The enzymes for the first two reactions, phosphoglucosamine mutase and glucosamine-1-phosphate acetyltransferase, as well as their genes, remain to be characterized. The N-acetylglucosamine-1-phosphate (GlcNAc-1-P) uridyltransferase activity (also named UDP-GlcNAc pyrophosphorylase), which synthesizes UDP-GlcNAc from GlcNAc-1-P and UTP, has previously been partially purified and characterized for Bacillus lichenifonnis and Staphylococcus aureus (1,34), * Corresponding author. but no data were available on the E. coli enzyme and its gene.…”

mentioning

confidence: 99%

Identification of the glmU gene encoding N-acetylglucosamine-1-phosphate uridyltransferase in Escherichia coli

1993

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The physiological properties of the EcoURF-1 open reading frame, which precedes the gbnS gene at 84 min on the Escherichia coli chromosome (J. E. Walker, N. J. Gay, M. Saraste, and A. N. Eberle, Biochem. J. 224:799-815, 1984), were investigated. A thermosensitive conditional mutant in which the synthesis of the gene product was impaired at 43°C was constructed. The inactivation of the gene in exponentially growing cells rapidly inhibited peptidoglycan synthesis. As a result, various alterations of cell shape were observed, and cell lysis finally occurred when the peptidoglycan content was 37% lower than that of normally growing cells. Analysis of the pools of peptidoglycan precursors revealed a large accumulation of N-acetylglucosamine-lphosphate and the concomitant depletion of the pools of the seven peptidoglycan nucleotide precursors located downstream in the pathway, a result indicating that the mutational block was in the step leading from N-acetylglucosamine-l-phosphate and UTP to the formation of UDP-N-acetylglucosamine. In vitro assays showed that the overexpression of this gene in E. col cells, directed by appropriate plasmids, led to a high overproduction (from 25-to 410-fold) of N-acetylglucosamine-l-phosphate uridyltransferase activity. This allowed us to purify this enzyme to homogeneity in only two chromatographic steps. The gene for this enzyme, which is essential for peptidoglycan and lipolysaccharide biosyntheses, was designated glnmU.

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“…Bacteria are also known to synthesize the cell-wall components peptidoglycan and lipopolysaccharide using UDP-GlcNAc derived from the well-known hexosamine biosynthesis pathway [1,14]. Three enzymes: (1) glucosamine-6-phosphate (GlcN-6-P) synthase encoded by glmS [4], (2) phosphoglucosamine mutase encoded by glmM [16], and (3) the unique bifunctional GlcN-1-P acetyltransferase and GlcNAc-1-P uridyltransferase encoded by glmU [15] have already been documented in Escherichia coli.…”

Section: Introductionmentioning

confidence: 99%

Acetate-mediated pH-stat fed-batch cultivation of transconjugant Enterobacter sp. BL-2S over-expressing glmS gene for excretive production of microbial polyglucosamine PGB-1

Son

Hong

Lee

2007

J Ind Microbiol Biotechnol

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A unique cationic polyglucosamine biopolymer PGB-1 comprising more than 95% D-glucosamine was excretively produced from a new bacterial strain Enterobacter sp. BL-2 under acetate-mediated culture conditions. Since the biopolymer PGB-1 could be synthesized from the UDP-N-acetylglucosamine monomer derived from the hexosamine pathway, three glmS, glmM, and glmU genes in the hexosamine pathway were cloned from Enterobacter sp. BL-2, and their molecular structures were elucidated. The cloned glmS, glmM, and glmU genes were reintroduced into the parent strain Enterobacter sp. BL-2 through a conjugative transformation for the overproduction of the biopolymer PGB-1. The biopolymer production increased 1.5-fold in the transconjugant Enterobacter sp. BL-2S over-expressing the first-step glmS gene encoding glucosamine-6-phosphate synthase. The transconjugant Enterobacter sp. BL-2S was cultivated pH-stat fed-batch widely, while intermittently feeding an acetate solution to maintain a constant pH level of 8.0 for 72 h, resulting in 1.15 g/L of the extracellular polyglucosamine biopolymer PGB-1.

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The control of synthesis of bacterial cell walls. Interaction in the synthesis of nucleotide precursors

Cited by 26 publications

References 19 publications

A conditional-lethal mutant of Bacillus subtilis 168 with a thermosensitive glycerol-3-phosphate cytidylyltransferase, an enzyme specific for the synthesis of the major wall teichoic acid

A conditional-lethal mutant of Bacillus subtilis 168 with a thermosensitive glycerol-3-phosphate cytidylyltransferase, an enzyme specific for the synthesis of the major wall teichoic acid

Identification of the glmU gene encoding N-acetylglucosamine-1-phosphate uridyltransferase in Escherichia coli

Acetate-mediated pH-stat fed-batch cultivation of transconjugant Enterobacter sp. BL-2S over-expressing glmS gene for excretive production of microbial polyglucosamine PGB-1

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