• Journal of Microbiology
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• 제33권4호
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• pp.344-349
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• 1995

The alkalophilic bacterium, Xanthomonas sp. JK311, producing extracellular proteases, was isolated from soil. Xanthomonas sp. JK311 produced five extracellular proteases that are all metalloproteases. Four of them were resistant against 1% SDS. Chromosomal DNA of the Xanthomonas sp. JK311 was digested with BamHI and cloned into PUC19. Among E. coli strain HB101 transformants, a clone secreting the proteases was screened through halo formation on skim-milk agar plate and by Southern blot analysis. It had the recombinant plasmid pXEP-1 containing the 7.5 kb-BamHI DNA fragment and produced three extacellular proteases. Their protease properties corresponded to those of Xanthomonas sp. JK311.

• Journal of Microbiology and Biotechnology
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• 제8권3호
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• pp.277-279
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• 1998

An extracellular polymer (exo-polymer) with hepatoprotective properties was produced after a 6-day submerged mycelial culture of Ganoderma lucidum WK-003. The glutamic pyruvic transaminase (GPT) activities in the serum of intoxicated Sprague-Dawley rats were decreased from 871 to 263 by the oral administration of the exo-polymer (20 mg/kg/day) for 4 consecutive days. Rhamnose, arabinose, xylose, mannose, galactose, and glucose were found in the exo-polymer along with aspartic acid, glutamic acid, histidine, serine, glycine, arginine, alanine, tryptophan, valine, phenylalanine, isoleucine, and leucine.

• KSBB Journal
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• 제23권1호
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• pp.23-30
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• 2008

An extracellular pigment production of three Phellinus sp. (Phellinus 421, P. linteus and P. hartigil) through submerged cultivation was investigated. The maximum brown pigment from culture broth was obtained from the precipitate by addition of 10% 1M HCI solution. This precipitate showed absorption characteristics with ${\lambda}_{max}$ of 360nm. The maximum production of extracellular pigment obtained at optimum medium and culture condition was 3.54 ($A_{360}$). The precipitate was fractionated by Sephadex G-75 gel chromatography, and the isolated brown pigment contained a large amount of polyphenol and the small amounts of sugar and protein. The brown pigment fraction was stable in temperature range of $30{\sim}60^{\circ}C$, pH range of $4{\sim}6$, sugar addition ranges of $1{\sim}5%$ and salt addition concentration of 3 molarity. Antioxidative activity of the brown pigment by TBA method was better than that of vitamin E (${\alpha}$-tocopherol).

• Applied Biological Chemistry
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• 제31권2호
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• pp.187-192
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• 1988

미생물에 의한 단백질 생산과 그의 분비기작에 관한 기초 자료를 얻고자 토양으로 부터 균체외 단백질 생산균주를 분리하고 동정한 결과 Bacillus sp.로 추정 되었다. 선정균주를 3.0%의 glucose와 1.3%의 urea를 첨가한 chemically defined medium에 접종하여 $30^{\circ}C$에서 48시간 배양하였을때 1.2mg/ml의 단백질을 생산 하였고 $CaCO_3$의 첨가로 단백질 생산이 촉진 되었으며 $250{\mu}g/ml$의 cephalexin를 첨가하여 96시간 배양 하였을때 2.0mg/ml의 단백질을 생산 하였다.

• Journal of Microbiology and Biotechnology
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• 제18권6호
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• pp.1136-1140
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• 2008

The Rhodopseudomonas palustris KUGB306 hemA gene codes for 5-aminolevulinic acid (ALA) synthase. This enzyme catalyzes the condensation of glycine and succinyl-CoA to yield ALA in the presence of the cofactor pyridoxal 5'-phosphate. The R. palustris KUGB306 hemA gene in the pGEX-KG vector system was transformed into Escherichia coli BL21. The effects of physiological factors on the extracellular production of ALA by the recombinant E. coli were studied. Terrific Broth (TB) medium resulted in significantly higher cell growth and ALA production than did Luria-Bertani (LB) medium. ALA production was significantly enhanced by the addition of succinate together with glycine in the medium. Maximal ALA production (2.5 g/l) was observed upon the addition of D-glucose as an ALA dehydratase inhibitor in the late-log culture phase. Based on the results obtained from the shake-flask cultures, fermentation was carried out using the recombinant E. coli in TB medium, with the initial addition of 90 mM glycine and 120 mM succinate, and the addition of 45 mM D-glucose in the late-log phase. The extracellular production of ALA was also influenced by the pH of the culture broth. We maintained a pH of 6.5 in the fermenter throughout the culture process, achieving the maximal levels of extracellular ALA production (5.15 g/l, 39.3 mM).

• Journal of Microbiology and Biotechnology
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• 제15권5호
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• pp.1054-1059
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• 2005

An extracellular protease was identified from Bacillus subtilis KCCM 10257 by N-terminal sequencing and mass spectral analysis. The molecular mass of the extracellular protease was estimated to be 28 kDa by SDS-PAGE. Sequencing of the N-terminal of the protease revealed the sequence of A(G,S,R)QXVPYG(A)V(P,L)SQ. The N-terminal sequence exhibited close similarity to the sequence of other proteases from Bacillus sp. A mass list of the monoisotopic peaks in the MALDI-TOF spectrum was searched after peptide fragmentation of the protease. Six peptide sequences exhibiting monoisotopic masses of 1,276.61, 1,513.67, 1,652.81, 1,661.83, 1,252.61, and 1,033.46 were observed from the fragmented protease. These monisotopic masses corresponded to the lytic enzyme L27 from Bacillus subtilis 168, and the Mowse score was found to be 75. A doubly charged Top product (MS) at a m/z of 517.3 exhibiting a molecular mass of 1034.6 was further analyzed by de novo sequencing using a PE Sciex QSTAR Hybrid Quadropole-TOF (MS/MS) mass spectrometer. MS/MS spectra of the Top product (MS) at a m/z of 517.3 obtained from the fragmented peptide mixture of protease with Q-star contained the b-ion series of 114.2, 171.2, 286.2, 357.2, 504.2, 667.4, 830.1, and 887.1 and y-ion series of 147.5, 204.2, 367.2, 530.3, 677.4, 748.4, 863.4, and 920.5. The sequence of analyzed peptide ion was identified as LGDAFYYG from the b- and y-ion series by de novo sequencing and corresponded to the results from the MALDI-TOF spectrum. From these results the extracellular protease from Bacillus subtilis KCCM 10257 was successfully identified with the lytic enzyme L27 from Bacillus subtilis 168.

• Archives of Pharmacal Research
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• 제29권11호
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• pp.1032-1041
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• 2006

Extracellular adenosine 5'-triphosphate (ATP) affects the function of many tissues and cells. To confirm the biological activity of ATP on human myeloid leukemic cells, F-36P and HL-60, cells were treated with a variety of concentrations of ATP. The stimulation with extracellular ATP induced the arrest of cell proliferation and cell death. from the analysis of Annexin-V staining and caspase activity by flow cytometry. The Annexin-V positive cells in both cell lines were dramatically increased following ATP stimulation. The expression of P2 purinergic receptor genes was confirmed, such as P2X1, P2X4, P2X5, P2X7 and P2Y1, P2Y2, P2Y4, P2Y5, P2Y6, P2Y11 in both leukemic cell lines. Interestingly, ATP induced intracellular calcium flux in HL-60 cells but not in F-36P cells, as determined by Fluo-3 AM staining. Cell cycle analysis revealed that ATP treatment arrested both F-36P and HL-60 cells at G1/G0. Taken together, these data showed that extracellular ATP via P2 receptor genes was involved in the cell proliferation and survival in human myeloid leukemic cells, HL-60 and F-36P cells by the induction of apoptosis and control of cell cycle. Our data suggest that treatment with extracellular nucleotides may be a novel and powerful therapeutic avenue for myeloid leukemic disease.

• Journal of Microbiology and Biotechnology
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• 제5권5호
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• pp.274-279
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• 1995

Ethyl caproate, one of the major flavor compounds deciding the quality of sake (Japanese wine), is produced during the brewing by the action of alcohol acyltransferase and esterases of sake yeast and koji mold. Extracellular esterases of Aspergillus oryzae required for ethyl caproate synthesis were purified partially. The enzymes had different optimum pH and affinity toward substrates. Substrate preferences and inhibition features showed the three enzymes to be B-type esterases or carboxylesterases (EC 3.1.1.1).

• Natural Product Sciences
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• 제18권4호
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• pp.211-214
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• 2012

Since 15-hydroxyprostaglandin dehydrogenase (15-PGDH) is the key metabolic enzyme of prostaglandin $E_2$ ($PGE_2$), inhibition of 15-PGDH is supposed to facilitate various physiological functions by increasing $PGE_2$. Methanol extract of Artemisia argyi (AAME) inhibited 15-PGDH ($IC_{50}$: $13.13{\mu}g/mL$) with relatively low cytotoxicity ($IC_{50}$: $415.00{\mu}g/mL$) and elevated extracellular $PGE_2$ levels in HaCaT cells. Real-time PCR analysis showed that AAME decreased significantly mRNA expression of PG transporter (PGT) in HaCaT cells. These results indicate that AAME could be applicable to functional materials as a 15-PGDH inhibitor and PGT expression inhibitor for the upregulation of extracellular $PGE_2$ level.

• Archives of Pharmacal Research
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• 제19권2호
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• pp.132-136
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• 1996

Exogenously applied oxygen free radical generating agent, pyrogallol, highly elevated extracellular glutamate accumulation and augmented N-methyl-D-aspartate (NMDA)-induced glutamate accumulation in cerebellar granule neuronal cells, but did not in astrocytes. Superoxide dismutase remarkably decreased the pyrogallol-induced glutamate accumulation, but either NMDA or kainate antagonists did not. In addition, pyrogallol did not affect the NMDAinduced intracellular calcium elevation. Pyrogallol partially blocked glutamate uptake into astrocytes. These results suggest that oxygen free radicals elevate extracellular glutamate accumulation by stimulating the release of glutamate as well as blocking the glutamate uptake.