• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.180-180
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• 1994

Farnesyl protein transferase는 Ras precursor의 C-terminal에 있는 cystein residue에 farnesyl group을 결합시키는 효소다. 이 효소를 bovine testis에서 30-50% ammonium sulfate fractionation, DEAE-sephacel ion exchange, Sephacryl s-300 gel filtration, hexapeptide(KKCVIM) affinity chromatography를 통해 30000배로 분리하였다. 분리된 효소는 gel filtration시 약 100kDa으로, SDS-polyacrylamide 전기영동시 50kDa의 인접한 두 bands로 나타났고 이것은 $\alpha$, $\beta$ subunits으로 생각되었다. $\alpha$ subunit을 encoding하는 RAM2 유전자를 site directed mutagenesis로 145번의 histidine을 aspartate로, 140번의 aspartate를 asparagine 으로 바꾸었더니 optimal pH와 $K_{m}$ 값이 변했다. Diethyl pyrocarbonate로 histidine residues를 chemical modification시켰을때 효소의 활성이 저하되었다. 145번 histidine이 aspartate로 바뀐 돌연변이효소에서 비교적 느리게 활성이 저하되므로 145번 histidine이 이 효소의 active site에 있을것으로 추측된다.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 1986.12a
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• pp.525.2-525
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• 1986

Cephalexin synthesizing enzyme (${\alpha}$ amino acid ester hydrolase) was partially purified from the culture broth of Acetobacter turbidans ATCC9325 through ammonium sulfate fractionation, DEAE, CM, and Sephacryl S-200 gel filtration. The enzyme has optimum pH 6.0 and temperature, 40$^{\circ}C$ respectively. From the analysis of reaction mixtures by thin layer chromatographic and high performance liquid chromatographic techniques, it was confirmed this enzyme catalyzed simultaneously the following reactions : 1) Synthesis of cephalexin from D-${\alpha}$-phenylglycine methylester (PGM) and 7-amino 3-deacetoxy-cetoxycephalosporanic acid (7-ADCA) 2) Hydrolysis of cephalexin to form 7-ADCA and phenylglycine (PG) 3) Hydrolysis of PGM to form PG and methanol. Base on the above experimental observations, the reaction model of this enzyme was identical with that of the enzyme from Xanthomonas citri.

• Microbiology and Biotechnology Letters
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• v.25 no.3
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• pp.271-276
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• 1997

An ascorbic acid phosphorylating enzyme, which catalyzes the formation of ascorbic acid-2-phosphate from ascorbic acid and pyrophosphate, was purified 32.7-folds to homogeneity from a cell-free extract of Cellulomonas sp. AP-7. The combination of DEAE- Sephacel ion exchange chromatography and Sephacryl S-200 get filtration was used for their purification. The molecular weight of the native protein was estimated to be 96.lkDa on high performance gel filtration chromatography. The SDS-PAGE analysis indicated that the protein consisted of four identical subunits of 24.6 kDa. The purified enzyme showed the optimal tempeature of 40$\circ$C and optimal pH of 4.5. The Km for ascorbic acid and pyrophosphate were 119 mM and 11.9 mM, respectively. The addition of 5,5'-dithiobis-(2-nitrobenzoic acid) into the reaction mixture resulted in the reduction of the enzyme activity at 51%. The enzyme also had a phosphatase activity at weakly acidic pH and the Km for ascorbic acid-2-phosphate in phosphatase activity was 7.9 mM.

• Journal of Applied Biological Chemistry
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• v.48 no.2
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• pp.75-78
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• 2005

Halotolerant protease produced by Halomonas marisflava KCCM 10457 was partially purified through ammonium sulfate precipitation and Sephacryl S-200HR gel permeation chromatography. Optimal pH and temperature of protease were 11.0 and $45^{\circ}C$, respectively. Enzyme activity was inhibited by $Cu^{2+}$, $Hg^{2+}$, $Fe^{2+}$, and $Fe^{3+}$, and selectively inhibited by p-chloromercuribenzoic acid (PCMB), suggesting this enzyme is cysteine protease. The enzyme is halotolerant, because it retained 77% of original activity in presence of 3.33 M NaCl. The protease showed broad substrate specificity to various natural proteins; BSA, casein, egg albumin, gelatin, and hemoglobin.

• Journal of Applied Biological Chemistry
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• v.54 no.2
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• pp.84-88
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• 2011

Edestin, a major hempseed storage protein from Cheungsam, was isolated to apparent homogeneity by acid precipitation and gel filtration chromatography. The native molecular weight of purified edestin was approximately 300 kDa by Sephacryl S-300 gel filtration. Upon adding 2-mercaptoethanol, the isolated edestin of 56.7 kDa on the non-reduced sodium dodecyl sulfate polyacrylamide gel was converted into subunits, suggesting that the protein might be composed of subunits linked by disulfide bond. Cheungsam edestin was rich in essential amino acids and it has 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity. The results suggest that Cheungsam edestin might be utilized as a superior antioxidative nutrient.

• Microbiology and Biotechnology Letters
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• v.17 no.6
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• pp.587-592
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• 1989

A strain of alkalophilic Bacillus sp. YS-309 capable of producing large amount of $\beta$-galactosidase has been isolated from soil sample. Intracellular $\beta$-galactosidase was purified 6.9 folds by procedures including ammonium sulfate precipitation, DEAE-cellulose chromatography, gel-filtration, DEAE-Sephadex A-50 chromatography with over-all yield of 17.8%. The molecular weight of native enzyme was 205, 000 by HPLC, and SDS-polyacrylamide gel electrophoresis showed that the enzyme consisted of 4 identical subunits with a molecular weight of 56, 000.

• Proceedings of the PSK Conference
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• 2003.04a
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• pp.217.3-218
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• 2003

A novel lectin has been purified from the fruiting bodies of the mushroom, Fomitella fraxinea, which belongs to bracket fungi by a combination of ion-exchange chromatography on DEAE-cellulose and gel filtration chromatography on Sephacryl S-200 HR. The lectin, designated as FFrL, was a homotetrametric protein with a molecular weight of 50 kDa as demonstrated by SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) and MALDI-TOF-MS(matrix assisted UV laser desorption/ionization time-of flight mass spectrometry). (omitted)

• Biotechnology and Bioprocess Engineering:BBE
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• v.9 no.5
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• pp.379-382
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• 2004

Entomopathogenic nematodes are used for insect control. Herein, an extracellular protease was partially purified from a culture supernatant of Xenorhabdus nematophilus, a symbiotic bacterium of an entomopathogenic nematode, Steinernema glaseri: using precipitation with $80\%$ v/v isopropyl alcohol followed by gel permeation chromatography with a packed Sephacryl S-300 HR media. The partially purified protease exhibited maximal activity at pH 7 in the presence of 1 mM $CaCl_2$. The protease was identified as a metallo-protease based on the inhibition of its activity by the metal chelating agent, EDTA.

• Archives of Pharmacal Research
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• v.14 no.3
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• pp.255-260
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• 1991

Chitinase was partially purified from Allium fistulosum L (green onion_. Protein fraction precipitated from ammonium sulfate was passed through CM-Sepharose and Sephacryl HR-200. The specific activity of the chitinase was 6.4 units/mg and total recovery was 6.3%. The analysis of the products from the digestion of N-acetychitohexaose indicated that chitinase was endo in action, with oligerms from N-acetylchitobiose to chitotetraose. N-Acetylglucosaminidase from the same species hydrolyzed oligomers obtained from chitinase reaction to lower oligosaccharides. These data demonstrated that chitinolytic system exists in green onion.

• Korean Journal Plant Pathology
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• v.14 no.3
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• pp.203-208
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• 1998

We isolated and characterized an antifungal peptide from the seeds of Phytolacca americana. Growth inhibition assay with Botrytis cinerea was used to screen inhibitory proteins from 60 different plant species. A 4 kDa antifungal peptide (Pa-AFP) inhibitory to hyphal growth of B. cinerea was found in the seeds of P. americana. The peptide Pa-AFP was purified to homogeneity by chromatographies of Sephadex G-50, DEAE-Sepharose, Sephacryl S-300, and C18 reverse-phase HPLC. Western blot analysis showed that a polyclonal antibody raised against the purified peptide cross-reacted with a 4 kDa protein in seeds but not in root and leaf tissues of P. americana. Pa-AFP inhibited the hyphal growth of Botrytis cinerea, Rihzoctonia solani, Fusarium oxysporum, and Magnaporthe grisea. Pa-AFP exhibited growth inhibition of Saccharomyces cerevisiae strain BWG7a, which was sensitive to osmotin.