• Journal of Microbiology and Biotechnology
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• 제15권5호
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• pp.919-923
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• 2005

A bacterium capable of poly(${\gamma}$-glutamic acid) production was isolated from nonpasteurized soy sauce. It was judged to be a variety of Bacillus subtilis and designated as B. subtilis C1. B. subtilis C1 produced ${\gamma}$-PGA in the absence of exogenous glutamic acid; therefore, it is a de novo PGA­producing bacterium. The product produced by B. subtilis C1 was characterized by amino acid analysis to be composed of solely glutamic acid. However, the $H^1-NMR$ spectra showed chemical shifts of glycerol protons in addition to those of authentic ${\gamma}$-PGA, indicating that the product is in fact a bioconjugate of ${\gamma}$-PGA. The finding is unique, because the microbial production of ${\gamma}$-PGA bioconjugate has never been reported before. The molecular mass of the product was over 10,000 kDa as determined by GPC, and $97\%$ of the product was D-glutamate, indicating that L-glutamate was converted to its D-form counterpart by B. subtilis C1.

• Nutrition Research and Practice
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• 제4권1호
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• pp.23-29
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• 2010

Poly-gamma-glutamic acid ($\gamma$-PGA) is a mucilaginous and biodegradable compound produced by Bacillus subtilis from fermented soybeans, and is found in the traditional Korean soy product, cheongkukjang. This study was carried out to evaluate the effects of $\gamma$-PGA from a food source on the concentration of the neurotransmitter GABA and its metabolic precursor glutamate in diet-induced obese rats. Eight-week old male Sprague-Dawley rats (n=60) were used. The rats were divided into two groups and obesity was induced by providing either a 10% control fat or 45% high fat diet for 5 weeks. The rats were then blocked into 6 groups and supplemented with a 0.1% $\gamma$-PGA diet for 4 weeks. After sacrifice, brain and serum GABA and glutamate concentrations were analyzed by high performance liquid chromatography with fluorometric detection. The rats fed the high fat diet had significantly increased body weights. $\gamma$-PGA supplementation significantly increased serum concentrations of glutamate and GABA in the control fat diet groups while this effect was not found in the high fat groups. In the brain, glutamate concentrations were significantly higher in the $\gamma$-PGA supplemented groups both in rats fed the normal and high fat diets than in the no $\gamma$-PGA controls. GABA concentrations showed the same tendency. The results indicated that $\gamma$-PGA intake increased GABA concentrations in the serum and brain. However, the effects were not shown in obese rats.

• Journal of Microbiology and Biotechnology
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• 제20권10호
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• pp.1436-1439
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• 2010

Many studies have clarified that poly(${\gamma}$-glutamic acid) (PGA) increases the solubility of $Ca^{2+}$, suggesting that PGA enhances calcium absorption in the small intestine. However, there has been no report on the specific interaction between PGA and $Ca^{2+}$ in water. We studied the aqueous solution properties of PGA calcium salt (PGA-Ca complex). The chelating ability and binding strength of PGA for $Ca^{2+}$ were evaluated. The PGA-Ca complex was soluble in water, in contrast to the insolubility of poly(acrylic acid) (PAA) calcium salt, and the chelating ability of PGA for $Ca^{2+}$ was almost the same as that of PAA. The globular conformation of the PGA-Ca complex in water was estimated by SEC and viscosity measurements. The chelation ability of PGA for $Ca^{2+}$ was examined by $^1H$ NMR. The present study showing the characteristics of the PGA-Ca complex will provide useful information about the calcium absorption by PGA in vivo.

• 한국식품영양과학회지
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• 제46권9호
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• pp.1114-1121
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• 2017

전통장류로부터 식품 유해요소를 생성하지 않는 Bacillus 균주를 분리하여 세포외효소 활성(amylase, protease, cellulase, xylanase)을 측정한 후, 단백질 분해 활성이 우수한 14개 균주와 비교균주 1균주를 선발하였다. 선발된 균주에 대해 16S rRNA 유전자를 이용한 균주 동정을 실시한 결과, B. amyloliquefaciens 10종, B. methylotrophicus 1종, B. velezensis 1종, B. subtilis 3종이 분리 동정되었다. 그중 B. subtilis JBG17019, B. amyloliquefaciens JBD17076, B. amyloliquefaciens JBD17109 균주에서 식중독미생물에 대한 증식 억제능이 확인되었다. Glutamic acid 대사와 관련한 발효특성을 확인하기 위하여 선발된 Bacillus 균주에 대해 glutamate, glutamine 및 ${\gamma}$-PGA 생성능을 측정하였다. 발효특성과 ${\gamma}$-PGA 생성능에 대한 다변량 요인분석을 주성분(PCA) 추출법으로 분석한 결과, PC1(효소 활성(amylase, cellulase, xylanase), PC2(${\gamma}$-PGA 생성능) 및 PC3(protease, glutamate 및 glutamine)의 3가지 주성분이 분류되었다. 주성분(PC)의 추출에 따라 B. amyloliquefaciens JBD17076 및 B. subtilis JBG17019 균주는 우수한 효소 활성 및 ${\gamma}$-PGA 생성을 하는 것으로 평가되었다.

• Journal of Microbiology and Biotechnology
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• 제15권1호
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• pp.59-65
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• 2005

Poly-${\gamma}$-glutamic acid (PGA) is an unusual anionic polypeptide and has great potential as an environmentally and industrially significant biodegradable material. A new ${\gamma}$-PGA producer, Bacillus mesentericus MJM1, with high production capacity was isolated from Korean domestic Chungkuckjang bean paste. It produced ${\gamma}$-PGA at the level of 10 g/l in suitable media. The viscosities of 5% initially extracted mucin and purified ${\gamma}$-PGA solutions were 660 cps and 600 cps, respectively. The produced ${\gamma}$-PGA polymer consisted of 2,000 glutamic acid residues with even proportion of L and D types with molecular mass of about 200- 300 kDa. Bacillus mesentericus MJM1 displayed ${\gamma}$-glutamyltranspeptidase (${\gamma}$-GTP) activity that is known to play a key role in ${\gamma}$-PGA biosynthesis. The ${\gamma}$-GTP coding region was located on the plasmid of 5.8 kb. The plasmid, named pMMH1, is a rolling-circle replication (RCR) plasmid and additionally contained a replication origin and type I signal peptidase (sipP) coding region.

• Biotechnology and Bioprocess Engineering:BBE
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• 제10권4호
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• pp.289-295
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• 2005

In this study, the cultural medium used for the efficient production of $\gamma$-PGA with a newly isolated Bacillus sp. RKY3 was optimized. It was necessary to supplement the culture medium with L-glutamic acid and an additional carbon source in order to induce the effective production of $\gamma$-PGA. The amount of $\gamma$-PGA increased with the addition of L-glutamic acid to the medium. The addition of 90 g/L L-glutamic acid to the medium resulted in the maximal yield of $\gamma$-PGA (83.2 g/L). The optimum nitrogen source was determined to be peptone, but corn steep liquor, a cheap nutrient, was also found to be effective for $\gamma$-PGA production. Both the $\gamma$-PGA production and cell growth increased rapidly with the addition of small amounts of $K_2HPO_4$ and $MgSO_4\cdot7H_{2}O$. Bacillus sp. RKY3 appears to require $Mg^{2+}$, rather than $Mn^{2+}$, for $\gamma$-PGA production, which is distinct from the production protocols associated with other, previously reported bacteria. Bacillus sp. RKY3 may also have contributed some minor $\gamma$-PGA depolymerase activity, resulting in the reduction of the molecular weight of the produced $\gamma$-PGA at the end of fermentation.

• Journal of Microbiology and Biotechnology
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• 제22권12호
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• pp.1782-1789
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• 2012

We have developed a novel type of polymer nanogel loaded with anticancer drug based on bio-derived poly(${\gamma}$-glutamic acid) (${\gamma}$-PGA). ${\gamma}$-PGA is a highly anionic polymer that is synthesized naturally by microbial species, most prominently in various bacilli, and has been shown to have excellent biocompatibility. Thiolated ${\gamma}$-PGA was synthesized by covalent coupling between the carboxyl groups of ${\gamma}$-PGA and the primary amine group of cysteamine. Doxorubicin (Dox)-loaded ${\gamma}$-PGA nanogels were fabricated using the following steps: (1) an ionic nanocomplex was formed between thiolated ${\gamma}$-PGA as the negative charge component, and Dox as the positive charge component; (2) addition of poly(ethylene glycol) (PEG) induced hydrogen-bond interactions between thiol groups of thiolated ${\gamma}$-PGA and hydroxyl groups of PEG, resulting in the nanocomplex; and (3) disulfide crosslinked ${\gamma}$-PGA nanogels were fabricated by ultrasonication. The average size and surface charge of Dox-loaded disulfide cross-linked ${\gamma}$-PGA nanogels in aqueous solution were $136.3{\pm}37.6$ nm and $-32.5{\pm}5.3$ mV, respectively. The loading amount of Dox was approximately 38.7 ${\mu}g$ per mg of ${\gamma}$-PGA nanogel. The Dox-loaded disulfide cross-linked ${\gamma}$-PGA nanogels showed controlled drug release behavior in the presence of reducing agents, glutathione (GSH) (1-10 mM). Through fluorescence microscopy and FACS, the cellular uptake of ${\gamma}$-PGA nanogels into breast cancer cells (MCF-7) was analyzed. The cytotoxic effect was evaluated using the MTT assay and was determined to be dependent on both the concentration and treatment time of ${\gamma}$-PGA nanogels. The bio-derived ${\gamma}$-PGA nanogels are expected to be a well-designed delivery carrier for controlled drug delivery applications.

• Journal of Microbiology and Biotechnology
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• 제27권5호
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• pp.1032-1037
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• 2017

The poly-${\gamma}$-$\small{D}$-glutamic acid (PGA) capsule, a major virulence factor of Bacillus anthracis, provides protection of the bacterium from phagocytosis and allows its unimpeded growth in the host. We investigated crosstalk between murine natural killer (NK) cells and macrophages stimulated with the PGA capsule of Bacillus licheniformis, a surrogate of the B. anthracis capsule. PGA induced interferon-gamma production from NK cells cultured with macrophages. This effect was dependent on macrophage-derived IL-12 and cell-cell contact interaction with macrophages through NK cell receptor NKG2D and its ligand RAE-1. The results showed that PGA could enhance NK cell activation by inducing IL-12 production in macrophages and a contact-dependent crosstalk with macrophages.

• Macromolecular Research
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• 제13권4호
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• pp.339-343
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• 2005

Microbial hydrogel was prepared by ${\gamma}-irradiation$ of poly(${\gamma}-glutamic acid$) (PGA) which was produced from Bacillus subtilis BS 62 and it's physico-chemical characteristic was examined. The hydrogel, prepared from 10% PGA with the dose of 48 kGy, was swollen up to 1,370 times of specific water content as dry weight basis. The hydrogels obtained above the dose of 48 kGy appeared to have higher compressive strength but lower specific water content. The period to reach a swelling equilibrium for the hydrogel in deionized water at the temperature range of 4 to $45^{\circ}C$ was about 10 h. The swollen hydrogel was shrunk in ionic solutions with the increase of ionic strength, and the rate of shrinkage was greater in calcium chloride solution than in sodium chloride. Specific water content of the hydrogel was quickly decreased at $80^{\circ}C$, showing a thennally hydrodegradable property.

• Journal of Microbiology and Biotechnology
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• 제29권9호
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• pp.1444-1452
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• 2019

The conventional prophylactic vaccines for human papillomavirus (HPV) efficiently prevent infection with high-risk HPV types, but they do not promote therapeutic effects against cervical cancer. Previously, we developed HPV16 E7-expressing Lactobacillus casei (L. casei-E7) as a therapeutic vaccine candidate for cervical cancer, which induces antitumor therapeutic effects in a TC-1 murine cancer model. To improve the therapeutic effect of L. casei-E7, we performed co-treatment with poly-gamma-glutamic acid (${\gamma}-PGA$), a safe and edible biomaterial naturally secreted by Bacillus subtilis. We investigated their synergistic effect to improve antitumor efficacy in a murine cancer model. The treatment with ${\gamma}-PGA$ did not show in vitro cytotoxicity against TC-1 tumor cells; however, an enhanced innate immune response including activation of dendritic cells was observed. Mice co-administered with ${\gamma}-PGA$ and L. casei-E7 showed significantly suppressed growth of TC-1 tumor cells and an increased survival rate in TC-1 mouse models compared to those of mice vaccinated with L. casei-E7 alone. The administration of ${\gamma}-PGA$ markedly enhanced the activation of natural killer (NK) cells but did not increase the E7-specific cytolytic activity of $CD8^+$ T lymphocytes in mice vaccinated with L. casei-E7. Overall, our results suggest that oral administration of ${\gamma}-PGA$ induces a synergistic antitumor effect in combination with L. casei-E7.