Expression of the<i>pgsB</i>Encoding the Poly-gamma-<scp>DL</scp>-glutamate Synthetase of<i>Bacillus subtilis (natto)</i>

Kimura, Keitarou; Tran, Lam‐Son Phan; Huyen, Thi; Itoh, Yoshifumi

doi:10.1271/bbb.80913

Cited by 38 publications

(24 citation statements)

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“…1). PgsB is the γPGA synthase dependent on ATP (Ashiuchi et al, 1999(Ashiuchi et al, , 2001Urushibata et al, 2002) and the pgsB gene is expressed in the early stationary phase (Kimura et al, 2009). These results suggested that the accumulation of ATP and/or an intermediate of γPGA are required for γPGA synthesis.…”

Section: Discussionmentioning

confidence: 99%

“…γPGA production is under the control of the ComQXPA quorumsensing system Tran et al, 2000) and the DegS DegU two-component system, and DegQ in particular plays an important role in γPGA production (Kimura et al, 2009;Ohsawa et al, 2009;Osera et al, 2009;Stanley and Lazazzera, 2005). Recently, SwrA, a protein associated with swarming motility, was reported to also be required for γPGA production (Osera et al, 2009;Stanley and Lazazzera, 2005).…”

Section: Introductionmentioning

confidence: 99%

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Disruption of the cell wall lytic enzyme CwlO affects the amount and molecular size of poly-γ-glutamic acid produced by Bacillus subtilis (natto)

Mitsui

Murasawa

Sekiguchi

2011

J. Gen. Appl. Microbiol.

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Poly-γ -glutamic acid (γ PGA), a polymer of glutamic acid, is a component of the viscosity substance of natto, a traditional Japanese food made from soybeans fermented with Bacillus subtilis (natto). Here we investigate the effects of the cell wall lytic enzymes belonging to the D,L-endopeptidases (LytE, LytF, CwlO and CwlS) on γ PGA production by B. subtilis (natto). γ PGA levels in a cwlO disruptant were about twofold higher than that of the wild-type strain, whereas disruption of the lytE, lytF and cwlS genes had little effect on γ PGA production. The molecular size of γPGA in the cwlO disruptant was larger than that of the wild-type strain. A complementary strain was constructed by insertion of the entire cwlO gene into the amyE locus of the CwlO mutant genome, and γ PGA production was restored to wild-type levels in this complementary strain. These results indicated that the peptidoglycan degradation enzyme, CwlO, plays an important role in γ PGA production and affects the molecular size of γ PGA. Key Words-Bacillus

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Disruption of the cell wall lytic enzyme CwlO affects the amount and molecular size of poly-γ-glutamic acid produced by Bacillus subtilis (natto)

Mitsui

Murasawa

Sekiguchi

2011

J. Gen. Appl. Microbiol.

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“…In B. subtilis, PGA secretion results in the production of a mucoid slime layer, whereas in Bacillus anthracis, PGA is retained to the cell surface as a capsule (20,21,22). In both B. subtilis and B. anthracis, an operon of conserved biosynthetic genes (pgsBCA or capABC genes, respectively) is both necessary and sufficient for PGA synthesis (23,24,25,26). In addition, B. subtilis encodes PgdS (also known as YwtD), which depolymerizes PGA (27,28).…”

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Defects in the Flagellar Motor Increase Synthesis of Poly-γ-Glutamate in Bacillus subtilis

2014

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Bacillus subtilis swims in liquid media and swarms over solid surfaces, and it encodes two sets of flagellar stator homologs. Here, we show that B. subtilis requires only the MotA/MotB stator during swarming motility and that the residues required for stator force generation are highly conserved from the Proteobacteria to the Firmicutes. We further find that mutants that abolish stator function also result in an overproduction of the extracellular polymer poly-␥-glutamate (PGA) to confer a mucoid colony phenotype. PGA overproduction appeared to be the result of an increase in the expression of the pgs operon that encodes genes for PGA synthesis. Transposon mutagenesis was conducted to identify insertions that abolished colony mucoidy and disruptions in known transcriptional regulators of PGA synthesis (Com and Deg two-component systems) as well as mutants defective in transcription-coupled DNA repair (Mfd)-reduced expression of the pgs operon. A final class of insertions disrupted proteins involved in the assembly of the flagellar filament (FliD, FliT, and FlgL), and these mutants did not reduce expression of the pgs operon, suggesting a second mechanism of PGA control.

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“…The functions of PgsA and the hydrophobic protein PgsC are ob-scure. The expression of the pgs operon is dependent on degQ, degU, and comA (13,30,32), suggesting that it is regulated by the cell density signaling pathway, as in the case of the degradation enzyme genes of the laboratory strain. However, how DegQ controlled ␥PGA synthesis was unclear (27,30).…”

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confidence: 99%

Mutations Suppressing the Loss of DegQ Function in Bacillus subtilis (natto) Poly-γ-Glutamate Synthesis

Huyen

Suzuki

Abe

et al. 2011

Appl Environ Microbiol

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The degQ gene of Bacillus subtilis (natto), encoding a small peptide of 46 amino acids, is essential for the synthesis of extracellular poly-gamma-glutamate (␥PGA). To elucidate the role of DegQ in ␥PGA synthesis, we knocked out the degQ gene in Bacillus subtilis (natto) and screened for suppressor mutations that restored ␥PGA synthesis in the absence of DegQ. Suppressor mutations were found in degS, the receptor kinase gene of the DegS-DegU two-component system. Recombinant DegS-His 6 mutant proteins were expressed in Escherichia coli cells and subjected to an in vitro phosphorylation assay. Compared with the wild type, mutant DegS-His 6 proteins showed higher levels of autophosphorylation (R208Q, M195I, L248F, and D250N), reduced autodephosphorylation (D250N), reduced phosphatase activity toward DegU, or a reduced ability to stimulate the autodephosphorylation activity of DegU (R208Q, D249G, M195I, L248F, and D250N) and stabilized DegU in the phosphorylated form. These mutant DegS proteins mimic the effect of DegQ on wild-type DegSU in vitro. Interestingly, DegQ stabilizes phosphorylated DegS only in the presence of DegU, indicating a complex interaction of these three proteins.Extracellular poly-gamma-glutamate (␥PGA) is a glutamate polymer linked through a gamma-peptide bond produced outside the cells by bacilli, including Bacillus subtilis, B. licheniformis, and B. anthracis, and Staphylococcus epidermidis (4). It acts as a nutrient reservoir to prevent starvation in the stationary phase and as a barrier against bacteriophage or phagocytotic attack by the host immune system (4, 12, 14, 16). Microbial functions that are not essential for the life cycle under laboratory conditions but that are required for survival in the natural environment are sometimes lost in domesticated laboratory strains. The loss of the ability to synthesize ␥PGA in a laboratory strain of B. subtilis is an example of such an event, and a mutation in degQ is involved in it (30).degQ function was characterized mainly through studies of the ␥PGA-negative laboratory lineage B. subtilis 168. degQ encodes a small peptide of 46 amino acids, and it is a pleiotropic regulator of degradation enzymes, including alkaline protease, ␣-amylase, and levansucrase (1). The expression of degQ is dependent on comA, the global transcriptional regulator in the stationary phase, and is strongly induced under dense-cell conditions (18). The regulation of these exoenzymes by degQ requires the presence of the degS-degU operon that comprises a two-component system (17, 18). The sensor histidine kinase DegS has four activities: it acts as a self-kinase (autophosphorylation of His189), a self-phosphatase (autodephosphorylation), a phosphotransferase (phosphorylation of Asp56 of DegU), and a phosphatase (dephosphorylation of DegU) (5,6,15,20,31). Exoenzyme expression is controlled directly by DegU, a DNA-binding protein, and DegU phosphorylation by DegS (DegU-P i ) enhances it (25,29,34). A recent study reported that DegQ stimulates phosphate transfer from phospho...

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Expression of thepgsBEncoding the Poly-gamma-DL-glutamate Synthetase ofBacillus subtilis (natto)

Cited by 38 publications

References 46 publications

Disruption of the cell wall lytic enzyme CwlO affects the amount and molecular size of poly-γ-glutamic acid produced by Bacillus subtilis (natto)

Disruption of the cell wall lytic enzyme CwlO affects the amount and molecular size of poly-γ-glutamic acid produced by Bacillus subtilis (natto)

Defects in the Flagellar Motor Increase Synthesis of Poly-γ-Glutamate in Bacillus subtilis

Mutations Suppressing the Loss of DegQ Function in Bacillus subtilis (natto) Poly-γ-Glutamate Synthesis

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