• Fisheries and Aquatic Sciences
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• 제12권4호
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• pp.249-257
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• 2009

This study is conducted to partially purify an antioxidative peptide in a two-step gelatin hydrolysate from Alaska pollock surimi refiner discharge, which was obtained by sequential treatment with Pronase E and Flavourzyme. The two-step gelatin hydrolysate was fractionated using chromatographic methods. Based on the same protein concentration of each fraction, the antioxidative activities (85.1-95.4%) of positive fractions fractionated by ion-exchange chromatography were higher than those (27.2-87.8%) from gel filtration. Then, further purification of the positive fractions was performed. Among them, the partially purified A1C1L2G1 and A1C1L2G2 fractions showed 96.2% and 85.1% inhibition, respectively, of linoleic acid peroxidation. The A1C1L2G1 fraction was composed of 15 kinds of amino acids and the predominant amino acids were proline, glycine and alanine. The results obtained in this study suggested that the fraction partially purified through chromatographic methods from the two-step gelatin hydrolysate of Alaska pollock surimi refiner discharge could be useful as a supplementary source for improving health functionality.

• 약학회지
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• 제41권5호
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• pp.615-621
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• 1997

Dehydropeptidase-I (DHP-I) was solubilized from porcine kidney by treatment with n-butanol and partially purified 19.25 fold by $(NH_4)_2SO_4$ precipitation, DEAE-Sepharose CL-6B ion exchange chromatography and Sephacryl S-300 HR chromatography with an overall yield of 19.16. DHP-I showed its optimal activity at pH 7.5 and 25$^{\circ}C$. Its activity was stable under neutral and alkaline conditions, but was disappeared under acidic condition. And DHP-I was heat-labile and its activity remained at 45$^{\circ}C$ for 3hrs. The enzyme was not inhibited by dicationic ions, while its activity was increased by $Co^{2+}$(1mM) and $Zn^{2+}$ (0.1mM). The enzyme was inhibited by EDTA and N-ethylmaleimide. The relative molecular mass of DHP-I was estimated to be approximately 100kDa by gel filtration chromatography. The $K_m$ value of DHP-I for glycyldehydrophenylalanine (GDHP) was 1.98mM. DWP20418 [(1R, 5S, 6S)-6-[1-(R)-Hydroxyethyl]-1-methyl-2-[(2S, 4S)-2-(piperazinylcarbonyl)-1-(R)-hydroxyethyl)pyrrolidine-4-thio]carbapen-2-em-3-carboxylic acid], compared with meropenem (MEPM), was rather easily hydrolized by DHP-I, while it was four times more resistant than imipenem (IPM) to DHP-I.

• 한국수산과학회지
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• 제54권6호
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• pp.927-937
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• 2021

This study was performed to screen the antimicrobial activities of the extract from the Pacific oyster Crassostrea gigas against skin pathogens and to purify the relevant antibacterial peptide. The acidified extract showed potent antibacterial activities against gram-positive and gram-negative bacteria but showed no activity against Candida albicans and no significant cell toxicity. Among acne-causing pathogens, the acidified extract showed potent antibacterial activity only against Staphylococcus aureus, and its antibacterial activity was completely abolished by treatment with trypsin or chymotrypsin, and was inhibited by salt treatment. The acidified extract showed strong DNA-binding ability but did not show bacterial membrane permeabilizing ability. Based on antimicrobial activity screening and cytotoxic effects, a novel antibacterial peptide was purified from the acidified gill extract using solid-phase extraction, cation-exchange, and reversed-phase HPLC. The resulting peptide had a molecular weight of 4800.8 Da and showed partial sequence homology with the carbonic anhydrase 4 (CA4) protein in the hard-shelled mussel. Overall, we purified a novel antibacterial peptide, named cgCAFLP, which is related to carbonic anhydrase 4 (CA4) protein, against skin pathogens. Our results suggest that the Pacific oyster extract could be used as an additive to control some acne-related skin pathogens (S. aureus).

• 한국수산과학회지
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• 제28권2호
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• pp.227-236
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• 1995

국내산 갈조류로부터 생리 기능성이 우수한 수용성 식이섬유 소재를 개발하기 위한 기초자료를 얻기위해 다시마와 미역(엽상체와 포자엽 각각)으로부터 fucoidan을 추출, 정제하여 그 특성을 조사하였다 Fucoidan의 추출 조건은 pH 2.0의 산성 추출액으로 $65^{\circ}C$I에서 1시간 씩 3회 추출을 하였고 cetylpyridinium chloride를 사용하여 부분 정제하였다. 부분 정제 fucoidan의 수율은 다시마 $2.71\%$, 미역포자엽 $6.65\%$, 미역 엽상체$0.40\%$였다. 특히 미역포자엽의 수율이 가장 높은 반면에 미역 엽상체는 가장 낮아 부위에 따른 차이가 매우 큼을 알 수 있었다. DEAE-Sephadex A-25 이온 교환 칼럼으로 부분 정제 fucoidan을 분획한 결과 다시마, 미역포자엽 모두 3종의 fraction으로 분획되었고 각 fraction은 전기 영동상에서 2종 이상의 band가 검출되어 불균일함을 알 수 있었다 이온 교환 칼럼으로 분획한 주fraction을 다시 알콜 분별 침전법으로 재분획한 결과 다시마와 미역포자엽은 에탄올농도 $60-70\%$에서 침전된 fraction이 cellulose acetate 전기 영동 및 gel filteration chromatography상에서 균일성을 나타내었다. 균일하게 정제된 fucoidan의 fucose : galactose : 황산기의 mole 조성비는 다시마, 미역포자엽이 각각 1 : 0.31 : 2.43,1 : 0.87 : 1.99였다. 분자량은 다시마가 31,000, 미역포자엽이 38,000으로 비교적 적었다.

• 생명과학회지
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• 제17권10호
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• pp.1321-1329
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• 2007

사슴뿔은 동물세계에서 가장 빨리 성장하는 조직이다. 따라서 성장중인 사슴뿔은 뼈 성장을 촉진하는 인자가 풍부하게 포함된 것으로 생각된다. 이들 성장인자들 중 IGF-1은 뼈를 자라게 하는 조골세포의 대사에 중요한 역할을 한다고 알려져 있어 이를 정제하고자 하였다. IGF-1의 정제는 상대라고 불리는 신선한 사슴뿔을 유안침전, DEAE-Sepharose CL-6B 이온교환수지, CM-Sepharose CL-6B 양이온교환수지, Sephadex G-50의 순차적인 방법으로 할 수 있었다. 각 과정마다 IGF-1의 거동을 HPLC, SDS-PAGE, Dot blot, 그리고 western blot으로 분석하였다. IGF-1의 정량은 ELISA기술로 재조합 인간 IGF-1을 이용하여 계산되었으며, 최종 분별 액은 두 개의 단백질을 보였으나, Western-blot에서 작은 분자량인 12 kDa으로 최종 판명할 수 있었다. 정제된 단백질은 HPLC에서 retention 시간 8분만에 검출되었으며, 총 농도는 2910 ng/ml 이고 중량은 0.291 g 이었다.

• Journal of Microbiology and Biotechnology
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• 제18권3호
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• pp.457-464
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• 2008

An extracellular enzyme (RMEBE) possessing ${\alpha}-(1{\rightarrow}4)-(1{\rightarrow}6)$-transferring activity was purified to homogeneity from Rhodothermus marin us by combination of ammonium sulfate precipitation, Q-Sepharose ion-exchange, and Superdex-200 gel filtration chromatographies, and preparative native polyacrylamide gel electrophoresis. The purified enzyme had an optimum pH of 6.0 and was highly thermostable with a maximal activity at $80^{\circ}C$. Its half-life was determined to be 73.7 and 16.7 min at 80 and $85^{\circ}C$, respectively. The enzyme was also halophilic and highly halotolerant up to about 2M NaCl, with a maximal activity at 0.5M. The substrate specificity of RMEBE suggested that it possesses partial characteristics of both glucan branching enzyme and neopullulanase. RMEBE clearly produced branched glucans from amylose, with partial ${\alpha}-(1{\rightarrow}4)$-hydrolysis of amylose and starch. At the same time, it hydrolyzed pullulan partly to panose, and exhibited ${\alpha}-(1{\rightarrow}4)-(1{\rightarrow}6)$-transferase activity for small maltooligosaccharides, producing disproportionated ${\alpha}-(1{\rightarrow}6)$-branched maltooligosaccharides. The enzyme preferred maltopentaose and maltohexaose to smaller maltooligosaccharides for production of longer branched products. Thus, the results suggest that RMEBE might be applied for production of branched oligosaccharides from small maltodextrins at high temperature or even at high salinity.

• Mycobiology
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• 제44권4호
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• pp.283-290
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• 2016

A double-stranded RNA (dsRNA) mycovirus was detected in malformed fruiting bodies of Pleurotus ostreatus strain ASI2792, one of bottle cultivated commercial strains of the edible oyster mushroom. The partial RNA-dependent RNA polymerase (RdRp) gene of the P. ostreatus ASI2792 mycovirus (PoV-ASI2792) was cloned, and a cDNA sequences alignment revealed that the sequence was identical to the RdRp gene of a known PoSV found in the P. ostreatus strain. To investigate the symptoms of PoV-ASI2792 infection by comparing the isogenic virus-free P. ostreatus strains with a virus-infected strain, isogenic virus-cured P. ostreatus strains were obtained by the mycelial fragmentation method for virus curing. The absence of virus was verified with gel electrophoresis after dsRNA-specific virus purification and Northern blot analysis using a partial RdRp cDNA of PoV-ASI2792. The growth rate and mycelial dry weight of virus-infected P. ostreatus strain with PoV-ASI2792 mycovirus were compared to those of three virus-free isogenic strains on 10 different media. The virus-cured strains showed distinctly higher mycelial growth rates and dry weights on all kinds of experimental culture media, with at least a 2.2-fold higher mycelial growth rate on mushroom complete media (MCM) and Hamada media, and a 2.7-fold higher mycelial dry weight on MCM and yeast-malt-glucose agar media than those of the virus-infected strain. These results suggest that the infection of PoV mycovirus has a deleterious effect on the vegetative growth of P. ostreatus.

• 산업식품공학
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• 제13권4호
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• pp.326-334
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• 2009

목이버섯으로부터 항혈전활성과 항혈소판응집활성을 지니는 물질을 추출하여 추출물의 혈행개선활성을 조사하였다. 건조 목이버섯을 0.1 N NaOH, methanol, ethanol 등의 용매를 사용하여 추출하여 각 추출물의 항혈전활성을 activated partial thromboplastin time, thrombin time과 prothrombin time 값으로 측정한 결과 methanol 용해성 분획이 각각 100, 124, 54 s로 조사한 분획 중에서 가장 높은 활성을 나타내었다. 활성 분획을 DEAE-Sepharose CL6B ion exchange column chromatography와 Sephacryl 400-HR gel permeation column chromatography로 정제하였으며 활성물질은 분자량이 150 kDa 이상인 다당류이며 mannose가 주요 구성당으로 되어 있는 xyloglucomannan의 복합다당체인 것으로 확인되었다. 정제된 다당류의 항혈전활성은 thrombin 활성을 저해하기 때문인 것으로 해석되었다.

• Journal of Microbiology
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• 제35권4호
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• pp.360-364
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• 1997

A ${\beta}$-ketothiolase was purified 180-fold from the cell extracts of Alcaligenes sp. SH-69 by a series of chromatography on DEAE-Dephadex A-50, Sephacryl S-200, and hydrozyapatitie columns, The optimum pH values of the partially purified enzyme were 7.5 for condensation reaction and 8.3 for thiolysis reaction were estimated to be 0.12mM and $18.7\;{\mu}M$, respectively. The $K_m$ valued for acetoacetyl-CoA and free CoASH in the thiolusis in the condensation reaction was 0.70mM. The condensation reaction of the ${\beta}$-ketothiolase was inhibited even by low concentrations of free CoASH($K_i=30.4{\mu}M$). Pretreatment of the enzyme with NADH and NADPH markedly inhibited the thiolysis reaction of the enzyme. The potent inhibition of the enzyme by sulfhydryl reagents suggests the involvement of cystein residue in the active site.

• Preventive Nutrition and Food Science
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• 제1권2호
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• pp.244-251
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• 1996

Molecular weight and partial amino acid sequence of the cis, 9-cis, 12-octadecadienoate isomerase(linoleate isomerase) of Butyrivibrio fibrisovens A-38 were determined. Linoleate isomerase was isolated from the bac-teria cultured anaerobically and purified by ultracentrifugation in conjunction with Sepharose 6B column chro-matography, Phenyl sepharose 4B column chromatography and fast performance liquid chromatography (EPLC). The isomerase was single polypeptide with 19KD of molecular weight, when determined by SDS-PAGE. Fourteen amino acids sequence of N-terminal of the linoleate isomerase was N-GEIDKYPRIIKQQ determined by Edman method.