Poly-γ-glutamate depolymerase of Bacillus subtilis: production, simple purification and substrate selectivity

Ashiuchi, Makoto; Nakamura, Hisaaki; Yamamoto, Takashi; Kamei, Tohru; Soda, Kenji; Park, Chung; Sung, Moon-Hee; Yagi, Toshiharu; Misono, Haruo

doi:10.1016/s1381-1177(03)00087-0

Cited by 41 publications

(34 citation statements)

References 22 publications

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“…Noteworthy, the pgs operons encode a ␥-glutamyl-hydrolase. This is the case for B. subtilis natto and B. licheniformis ATCC 14580, which possess the ␥-glutamylhydrolase genes pgsS and ywtD, respectively (2,36). The only well-characterized example of PGA-capsulated bacteria is B. anthracis, which possesses CapD, a ␥-glutamyltranspeptidase.…”

Section: Discussionmentioning

confidence: 99%

CapE, a 47-Amino-Acid Peptide, Is Necessary for Bacillus anthracis Polyglutamate Capsule Synthesis

2005

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Polyglutamate is found in various bacteria, but displays different functions depending on the species and their environment. Here, we describe a minimal polyglutamate synthesis system in Bacillus anthracis. In addition to the three genes previously described as sufficient for polyglutamate synthesis, this system includes a small open reading frame, capE, belonging to the cap operon. The polyglutamate system's requirement for the five cap genes, for capsulation and anchoring, was assayed in nonpolar mutants. The capA, capB, capC, and capE genes are all necessary and are sufficient for polyglutamate synthesis by B. anthracis. capD is required for polyglutamate anchoring to the peptidoglycan. The 47-amino-acid peptide encoded by capE is localized in the B. anthracis membrane. It is not a regulator and it is required for polyglutamate synthesis, suggesting that it has a structural role in polyglutamate synthesis. CapE appears to interact with CapA. Bacillus subtilis ywtC is similar to capE and we named it pgsE. Genes similar to capE or pgsE were found in B. subtilis natto, Bacillus licheniformis, and Staphylococcus epidermidis, species that produce polyglutamate. All the bacterial polyglutamate synthesis systems analyzed show a similar genetic organization and, we suggest, the same protein requirements.

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Section: Discussionmentioning

confidence: 99%

CapE, a 47-Amino-Acid Peptide, Is Necessary for Bacillus anthracis Polyglutamate Capsule Synthesis

2005

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“…The biosynthesis of DL-PGA by some strains of B. subtilis has been well studied (4,6,30), but both the substrate specificities of their PGA synthetic systems and the structural features of the reaction products remained obscure. Figure 5 shows the structural features of the PGAs synthesized by the PGA synthetic system of B. subtilis subsp.…”

Section: Vol 70 2004 Enzymatic Synthesis Of Poly-␥-glutamate 4251mentioning

confidence: 99%

“…To address the former problem, many attempts have been made to isolate and construct industrially useful producers of PGA (4,9,20). Considering the fact that DL-PGA from B. subtilis, which shows irregular stereochemistry (4,6,30), is currently available, the latter problem may be more profound. In general, the thermoplasticity of biodegradable polymers is significantly influenced by the homogeneity of the stereochemical compositions (12).…”

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confidence: 99%

Enzymatic Synthesis of High-Molecular-Mass Poly-γ-Glutamate and Regulation of Its Stereochemistry

Ashiuchi

Shimanouchi

Nakamura

et al. 2004

Appl Environ Microbiol

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For the first time, we succeeded in synthesizing in vitro poly-␥-glutamate (PGA) with high molecular masses (>1,000 kDa) by the use of enzyme-associated cell membranes from Bacillus subtilis subsp. chungkookjang. The activity for PGA synthesis, however, was readily lost in the presence of critical concentrations of detergents tested in micelles. The optimum pH for the reaction was found to be ϳ7.0. We examined the effects of some divalent cations on PGA synthesis and found that Mg 2؉ was essential in catalysis and that Zn 2؉ additionally boosted the activity. In contrast, Fe 2؉ and Ca 2؉ acted as inhibitors. Mn 2؉ did not apparently influence the in vitro formation of PGA. DL-Glutamate (D isomer content, 60 to 80%) apparently served as the best substrate; D-Glutamate was preferable to the L isomer as a substrate. When D-and L-glutamate were used for the reaction, the elongated chains of PGAs were composed of the D-and L-isomers, respectively. Our results suggest that the stereochemical properties of enzymatically synthesized PGAs substantially depend on the stereochemistry (DL ratio) of glutamate as the substrate. Furthermore, genetic analysis indicated that all the pgsB, -C, and -A gene products, which are responsible for PGA production by B. subtilis cells, were also indispensable for enzymatic PGA synthesis.

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“…c Stereochemical compositions of glutamate in PGA hydrolysates were assayed by a chiral high-performance liquid chromatography method (3)(4)(5).…”

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Salt-Inducible Bionylon Polymer from Bacillus megaterium

Shimizu

Nakamura

Ashiuchi

2007

Appl Environ Microbiol

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Poly-γ-glutamate depolymerase of Bacillus subtilis: production, simple purification and substrate selectivity

Cited by 41 publications

References 22 publications

CapE, a 47-Amino-Acid Peptide, Is Necessary for Bacillus anthracis Polyglutamate Capsule Synthesis

CapE, a 47-Amino-Acid Peptide, Is Necessary for Bacillus anthracis Polyglutamate Capsule Synthesis

Enzymatic Synthesis of High-Molecular-Mass Poly-γ-Glutamate and Regulation of Its Stereochemistry

Salt-Inducible Bionylon Polymer from Bacillus megaterium

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