• Journal of Microbiology and Biotechnology
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• 제21권1호
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• pp.20-27
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• 2011

The purification and characterization of a $Mn^{2+}$-dependent alkaline serine protease produced by Bacillus pumilus TMS55 were investigated. The enzyme was purified in three steps: concentrating the crude enzyme using ammonium sulfate precipitation, followed by gel filtration and cation-exchange chromatography. The purified protease had a molecular mass of approximately 35 kDa, was highly active over a broad pH range of 7.0 to 12.0, and remained stable over a pH range of 7.5 to 11.5. The optimum temperature for the enzyme activity was found to be $60^{\circ}C$. PMSF and AEBSF (1 mM) significantly inhibited the protease activity, indicating that the protease is a serine protease. $Mn^{2+}$ ions enhanced the activity and stability of the enzyme. In addition, the purified protease remained stable with oxidants ($H_2O_2$, 2%) and organic solvents (25%), such as benzene, hexane, and toluene. Therefore, these characteristics of the protease and its dehairing ability indicate its potential for a wide range of commercial applications.

• 대한화학회지
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• 제43권1호
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• pp.52-57
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• 1999

In($C_6F_5)_3{\cdot}D(D=CH_3CN$, O($C_2H_5)_2$)와 ($CH_3)_3SiC_6F_5$/CsF, $C_6F_5$MgBr 또는 Cd($C_6F_5)_2$을 반응시켜 [In($C_6F_5)_4$]- 음이온 화합물을 합성하였으나, 이들 indium(III) 음이온 화합물들은 온도에 민감하고 습기에 대해 불안정하다. 안정한 indium(III) 음이온 착물은 PNPCl(PNP=bis(triphenylphosphino)ammonium)과의 양이온 치환반응시켜 얻었으며, 관 크로마토 그래피를 이용하여 분리 정제하였다. 합성된 화합물의 특성은 핵자기 공명 분석법, 적외선 분광분석법, 분자량 측정, DTA/TG 그리고 원소분석법을 이용하여 조사하였다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제9권1호
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• pp.1-6
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• 2004

The human chemokine, the short version of leukotactin-1(shLkn-1;molecular weight=7.2 kD and 66 amino acids), was expressed and secreted into a culture medium using the me-thylotrophic yeast, Pichia pastoris. The recombinant shLkn-1 was purified from the culture supernatant using a simple two-step procedure consisting of cation exchange and reverse phase chromatography(RPC), in which shLkn-1 was highly purified (99.5%) with a high recovery yield of 82.7%. The C-terminal truncated derivative of shLkn-1 was found in the supernatant and was separated by RPC. The physicochemical properties of the purified shLkn-1 were verified to be the same as expected. The biological activity of the purified recombinant shLkn-1 was also quantified using a chemotaxis assay. It was observed that the recombinant shLkn-1 had the maximum migration activity at a concentration of 10nM, as potent as MIP-1${\alpha}$.

• 한국식품영양과학회지
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• 제3권1호
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• pp.69-76
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• 1974

The method of amino acid sequence determination from the C-terminal amino acid is proposed and mass spectrometric identification of thiohydantoins described previously. In this paper was discussed the fragmentation of thiohydantoin-ring by deutero substitution and model tripeptide have been degraded through three stages each, with interpretable results. The conditions employed in this method are mild enough for biological materials. The main features of the method are the following. 1. Thiohydantoins were formed in a non-aqueous medium a mixture of acetic anhydride, acetic acid and ammonium thiocyanate. 2. Mass sepectra of thiohydantoins derived from 20 amino acids were obtained with a mass spectrometer, JEOL model JMS-06H. 3. Cleavage of peptidyl thiohydantoin was made with an acidic from of a cation-exchange resin. (Amberlite IR-120) 4. Separation of the cleaved thiohydantoin and the parent peptide less one amino acid moiety was made by chromatography on a Sephadex G-10 column. 5. The peptide fraction was concentrated by freezedrying. 6. Thiohydantoin derivative of carboxyl terminal amino acid residue was introduced with a direct inlet probe in methanol solution.

• Nuclear Engineering and Technology
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• 제12권2호
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• pp.99-105
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• 1980

과망간산염, 염소산염및 과염소산염의 중성자 포획에 수반하는 망간-56 및 염소-38 반조화학종의 분배를 이온교환크로마토구라휘법으로 검토하였다. 과망산염의 망간-56 방사능은 2종의 원자가상태로 나타나며 염소산염의 염소-38은 2종의 원자가상태 그리고 과염소산염의 염소-38은 3종의 원자가상태로 나타난다. 반조에너지는 계산하였다. $^{38m}$Cl의 핵이성체 전이에 수반하는 내부전환은 잔류율에 영향을 미친다. 양이온반경이 클수록 2차 케이지를 돌파하는 반조원자의 확율이 높다. 암모늄염중 암모늄이온은 환원제 작용을 한다. 결정구조에서 자유공간이 크면 잔류율은 낮아진다.다.

• BMB Reports
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• 제31권4호
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• pp.328-332
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• 1998

The 8-hydroxyisocoumarin, named Centipedin, which has a significant antibiotic activity, was separated and solubilized with organic solvents, such as diethyl ether from centipede Scolopendra subspinipes multilans L. Koch. The Centipedin was purified by silicic acid column and high S cation exchange chromatography followed by reverse-phase HPLC. It was confirmed that Centipedin has a potent antibiotic effectiveness against Gram-negative Klebsiella pneumoniae ATCC 8308. The results showed that Centipedin blocks both DNA replication and RNA transcription during the growth of this pathogen in vivo. The biosynthesis of antibiotic 8-hydroxyisocoumarin was studied in vivo by feeding $[^{14}C]-labelled$ compound as a precursor to live centipede, in which $[^{14}C]acetate$ was the most efficiently incorporated into the Centipedin within 30 h after injection. Also, in vitro study on the biosynthesis of Centipedin showed that efficient incorporation of $[^{14}C]acetate$ occurred at pH range 5.0-7.0 for 10 h incubation and decreased significantly after then. It is suggested that 8-hydroxyisocoumarin is one of the defense compounds acting on bacterial infection in Scolopendra subspinipes.

• Journal of Plant Biotechnology
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• 제32권4호
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• pp.269-273
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• 2005

Methylotrophic 효모 Pichia pastoris 발현시스템을 사용하여 인간 TLR9 단백질의 세포내 TIR 도메인을 발현하였다. TIR 단백질이 P. pastoris에서 발현되어 배지 속으로 분비되는 것을 SDS-PAGE로 확인하였고, 발현된 단백질을 western-blot, MALDI-TOF 질량분석으로 동정하였다. 이를 통하여 TIR 딘백질이 P. pastoris에서 안정적으로 발현됨을 알 수 있었다. 그리고 발현된 단백질을 니켈 친화, 양이온교환수지, 겔 투과 크로마토그라피를 사용하여 순수 분리 정제하였다. P. pastoris를 이용한 단백질의 발현과 정제방법은 대장균에서 잘 발현되지 않는 단백질의 발현에 응용될 수 있을 것이다.

• Journal of Microbiology and Biotechnology
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• 제7권6호
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• pp.409-416
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• 1997

Leuconostoc sp. LAB145-3A isolated from kimchi produced a bacteriocin which was active against food pathogens, such as Listeria monocytogenes, Enterococcus faecalis, and E. faecium. Bacteriocin production occurred during the early exponential phase of growth and was stable upto the late stationary phase of growth. Optimum conditions for bacteriocin production were $37^{\circ}C$ with an initial pH of 7.0. The bacteriocin of LAB145-3A was sensitive to proteases, but stable for solvents, pH change and heat treatment. It was stable even at autoclaving temperature for 15 min. The bacteriocin exhibited a bactericidal mode of action against Lactobacillus curvatus LAB170-12. The bacteriocin produced by Leuconostoc sp. LAB145-3A was purified by CM-cellulose cation exchange column chromatography and Sephadex G-50 gel filtration. The purification resulted in an approximate 10,000-fold increase in the specific activity. Approximately 4% of the initial activity was recovered. Purified bacteriocin exhibited a single band on the SDS-PAGE with an apparent molecular weight of 4,400 daltons. This bacteriocin was named leucocin K. Leuconostoc sp. LAB145-3A had two residential plasmids with molecular sizes of 23 kb and 48 kb. A comparison of plasmid profiles between LAB145-3A and its mutants revealed that the 23 kb plasmid (pCA23) was responsible for bacteriocin production and immunity to the bacteriocin in Leuconostoc sp. LAB145-3A.

• 한국수산과학회지
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• 제49권2호
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• pp.137-145
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• 2016

This study screened the biological activity of an acidified gill extract of the Manila clam Ruditapes philippinarum including antimicrobial, hemolytic, membrane permeabilization, and DNA-binding activity, and purified the antimicrobial material. The acidified gill extract showed potent antimicrobial activity against Bacillus subtilis and Escherichia coli without significant hemolytic activity, but showed no membrane permeabilization or DNA-binding ability. An antimicrobial material was purified from the acidified gill extract using C₁₈ reversed-phase and cation-exchange high-performance liquid chromatography (HPLC). Treatment of the purified material with trypsin completely abolished all of the antibacterial activity against Bacillus subtilis, suggesting that the purified material is a proteinaceous antibiotic. The molecular weight of the purified material was 2571.9 Da, but no primary structural information was obtained due to N-terminal blocking. A future study should confirm the primary structure. Our results suggest that the Manila clam gill contains proteinaceous antibiotics that have a role in first-line defense. This information could be used to better understand the Manila clam innate immune system.

• Journal of Microbiology and Biotechnology
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• 제13권2호
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• pp.269-275
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• 2003

Chitosan-degrading activity induced by chitosan was founf in culture filtrate of Aspergillus flavus IAM2044. Aspergillus flavus IAM2044 had a higher level of chitosanolytic activity when chitosan was used as a carbon source, and yeast extract and peptone were supplemented as nitrogen sources. One of the chitosan-degrading enzymes was purified to homogeneity by ammonium sulfate precipitation followed by cation-exchange and gel filtration chromatographies. The enzyme was monomeric, and its molecular mass was 45 kDa. The optimum pH and temperature of the enzyme were 5.0 and $50^{\circ}C$, respectively. The activity was stable in the pH range of 3.5 to 7.0 and at a temperature below $50^{\circ}C$. Reaction products analyzed by the viscosimetric assay and thin layer chromatography clearly indicated that the enzyme was an exe-type chitosanase, $exo-{\beta}-D-glucosaminidase$, that released GlcN from the nonreducing ends of the oligosaccharide chains.