• BMB Reports
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• 제29권4호
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• pp.294-299
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• 1996

Glyoxalase I (Ee 4.4.1.5, lactoylglutathione lyase) from Chlamydomonas reinhardtii was purified to homogeneity by ammonium sulfate fractionation, anion-exchange chromatography, hydrophobic interaction chromatography, and affinity chromatography on S-hexylglutathione agarose. The purified enzyme was judged to be homogeneous on SDS-PAGE, and consisted of a single polypeptide chain with a relative molecular weight of 24,000. The enzyme was most active at $40^{\circ}C$ and pH 7.5. It was catalytically most active with methylglyoxal as substrate. A number of properties of the Chlamydomonas glyoxalase I enzyme, such as substrate specificity, molecular mass, kinetic parameters, pi, metal ion effect, have been determined and compared with those reported for preparations from other sources. It had somewhat different characteristics from mammalian enzymes.

• 한국생물물리학회:학술대회논문집
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• 한국생물물리학회 2002년도 제9회 학술 발표회 프로그램과 논문초록
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• pp.53-53
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• 2002

Inhibitory effects of Cu$\^$2＋/ on the cytochrome P450 (P450)-catalyzed reactions of liver microsomes and reconstituted systems containing purified P450 and NADPH-P450 reductase (NPR) were seen. However, Zn$\^$2＋/, Mg$\^$2＋/, Mn$\^$2＋/, Ca$\^$2＋/, and Co$\^$2＋/ had no apparent effects on the activities of microsomal P450s. Cu$\^$2＋/ inhibited the reactions catalyzed by purified P450s lA2 and 3A4 with IC$\sub$50/ values of 5.7 and 8.4 ${\mu}$M, respectively.(omitted)

• 미생물학회지
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• 제26권1호
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• pp.52-59
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• 1988

A partially purified preparation of L-galactonolactone oxidase which catalyzes the last step of L-ascorbic acid biosynthesis was obtained from Saccharomyces cerevisiae ATCc 26787. The purification procedures included Triton X-100 treatment, protamine sulfate precipitation, ammonium sulfate precipitation, DEAE-Sepharose CL-6B ion exchange chromatography, Sephadex G-150 gel filtration chromatography, and Phenyl-Sepharose CL-4B hydrophobic interaction chromatography. The optimum temperature for the enzyme activity was about $34^{\circ}C$ and the optimum pH was 6.8-7.0. The substrate specificity was confined to L-aldonolactones, L-galactono-1,4-lactone and L-gulono-1,4-lactone. An apparent Km value of 0.294mM with L-galactono-1,4-lactone as a substrate was found. By comparing the substrate specificities of this enzyme with those of isofunctional enzymes of higher plants and animals, it becomes evident that the enzyme of S. cerevisiae ATCC 26787 is rather similar to the L-gulonolactone oxidase of animals than the galactonolactone dehydrogenase of higher plants.

• Preventive Nutrition and Food Science
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• 제1권1호
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• pp.106-110
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• 1996

Bromelain(EC 3.4.22.4), the collective name for the proteolytic enzymes found in tissues of the plant family Bromeliaceae(pineapple), has been used as a tenderizing agent in food processing, and as an antiinflammatory agent in pharmaceuticals. In this paper, we describe the simple purification method of bromelain using Korean pineapple fruit. After removing contaminants at 30% saturation of ammonium sulfate, the supernatant obtained was treated again with ammonium sulfate to 80% saturation. Wit the above salt fractionation, partially purified bromelain could be obtained. To get highly purified bromelain, the previous 30% to 80% ammonium sulfate treated precipitate was dialyzed against 25mM sodium acetate buffer(pH 5.0) followed by passing through a CM- cellulose cation exchange column. Fruit bromelain was eluted as a major peak at 0.5~0.8M NaCI gradient. The present method is simpler with high wield than the traditional purification method-acetone treatment and several consecutive chromatographic processes.

• Journal of Microbiology and Biotechnology
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• 제11권6호
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• pp.946-950
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• 2001

Slow-growing pathogenic mycobacterium species, including Mycobacterium bovis BCG, secrete a large amount of glutamine synthetase into culture media. Extracellular and intracellular glutamine synthetases were purified from M. bovis BCG. While the native molecular weights of both glutamine synthetases were estimated to be 370.2 kDa, those of the subunits were 61.7 kDa, indicating that the native forms were composed of 6 subunits. The enzymes showed a hhigh thermal stability and high degree of sequence similarity with the glutamine synthetase from M. tuberculosis in the N-terminal amino acid sequence. Western blotting analysis indicated that the antibodies prepared against both the extracellular and intracellular enzymes exhibited common antigen determinants.

• 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).

• Journal of Microbiology and Biotechnology
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• 제1권3호
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• pp.176-181
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• 1991

Studies on the optimum conditions for dextran production and the properties of dextransucrase (DS) were performed with Leuconostoc mesenteroides from Sikhae and Leuconostoc mesenteroides NRRL B-512(F). Dextransucrases were partially purified by lyophilization of the culture supernatant and subsequent gel chromatography on Bio-Gel A-5(m). The storage stabilities of Sikhae DS and B-512(F) DS were decreaed by the addition of dextranase. The optimum conditions for the enzyme stability were pH 5 and below $30^{\circ}C$. The B-512(F) DS lost the activity at pH 4, while Sikhae DS had 30% of the activity at the same pH. The activity of DS was decreased by EDTA. confirming the metalloprotein character of the enzymes, and was restored by the addition of calcium ions. Concanavalin A completely removed the activity of DSs, confirming the glycoprotein character of the enzymes.

• 한국미생물·생명공학회지
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• 제25권2호
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• pp.189-196
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• 1997

A producer of inhibitor against ${\beta}-glucosidase$ of Penicillium sp.-L4 was screened from Actinomycetes, and the isolated strain was identified as Streptomyces sp. The inhibitor produced was very stable against heat, acidic and alkaline conditions, proteolytic and amylolytic enzymes. The inhibotor was purified from culture broth through activated carbon treatment, ultrafiltration, anion and cation exchange, activated carbon columm, acetone precipitation and preparative HPLC. It showed inhibitory activities against a variety of dissacharide hydrolyzing enzymes produced by P.sp.-L4, and the mode of inhibition was competitive. Its structure and molecular formular was elucidated by IR, $^1H\;and\;^{13}C$ NMR and FAB/Mass spectrometry, which was identified as 1-deoxynojirimycin (dNM). dNM showed inhibitory effects on the cell growth and hydrolytic enzyme action of P.sp.-L4 on agar plate and infected lemon peel.

• Molecules and Cells
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• 제25권2호
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• pp.312-316
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• 2008

Flavanone $3{\beta}$-hydroxylases (F3H) are key enzymes in the synthesis of flavonol and anthocyanin. In this study, three F3H cDNAs from Oryza sativa (OsF3H-1 ~3) were cloned by RT-PCR and expressed in E. coli as gluthatione S-transferase (GST) fusion proteins. The purified recombinant OsF3Hs used flavanone, naringenin and eriodictyol as substrates. The reaction products with naringen and eriodictyol were determined by nuclear magnetic resonance spectroscopy to be dihydrokaempferol and taxifolin, respectively. OsF3H-1 had the highest enzymatic activity whereas the overall expression of OsF3H-2 was highest in all tissues except seeds. Flavanone $3{\beta}$-hydroxylase could be a useful target for flavonoid metabolic engineering in rice.

• Mycobiology
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• 제29권4호
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• pp.183-189
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• 2001

Extracellular amylase properties were examined with the mycelium of Tricholoma matsutake isolated from ectomycorrhizal roots of Pinus densiflora. The molecular weights of $\alpha$-amylase and glucoamylase were estimated as 34.2 kD and 11.5 kD, respectively, after eluted through Superdex 75 column. The optimum pH of the purified enzyme was found in a range of pH $5.0{\sim}6.0$, with a peak at pH 5.0. The activities of these enzymes were stable from $4^{\circ}C\;to\;30^{\circ}C$. The $\alpha$-amylase of T. matsutake readily hydrolyzed soluble starch and amylose-B, while it weakly hydrolyzed glycogen, dextrin, amylose and amylose-A. The main products of hydrolysis were confirmed to be glucose, maltose and maltotriose on the basis of the similarities in the thin layer chromatographic mobility.