本体／０１‐近泉・中村・水谷

Size: px

Start display at page:

Download "本体／０１‐近泉・中村・水谷"

あきみにかどり
5 years ago
Views:

1 Mem. Fac. Agr., Ehime Univ. Saccharomyces cerevisiae Hong mei ZHAO, Yudai SHIRAISHI, Hiroki TANAKA and Yasuo WATANABE : Purification of Glycerol Dehydrogenases from Saccharomyces cerevisiae and the Substrate Specificity of Bacterial Enzymes Abstract We have obtained the dehydrogenases being specific for mannitol and arabitol with the gene cloning and over-expression in yeast cells, in order to measure the content of various sugar alcohols by using the dehydrogenases. In this study, we carried out the isolations of glycerol dehydrogenases from Saccharomyces cerevisiae and determined its substrate specificities. Target proteins were purified by affinity chromatography with the addition of tag at N-terminus or C-terminus by gene recombination. The obtained enzymes showed the low specificity for sugar alcohols. However, we found the bacterial enzymes that showed the high specificity for glycerol in commercial products. These glycerol dehydrogenases could be available in the enzymatic analysis of glycerol content. polyhydroxyalkane D D DD β Onishi van Zyl and Prior D D D foods/index.html Onishi van Zyl and Prior Onishi Iwaki et al.watanabe et al. Watanabe et al.zygosaccharomyces rouxii Saccharomyces cerevisiae Iwaki et al.z. rouxii van Zyl et

2 al. Tang et al. de Figueroa and Lucca S. cerevisiae ORF YEL070w Watanabe et al. Minamihara and Suzuki Kluyveromyces lactis D S. cerevisiae Dujon et al. Gelperin et al. S. cerevisiae GCY1 YPR1 N C S. cerevisiae GCY1 YPR1 ORF BG His peghs ORF S S-Powder - ura Difco g g Ile g Val g Ade g Arg g His g Leu g Lysg Met g Phe g Trp g Tyr g SC ura SC - ura ml S-Powder - ura g g SC - ura ml S-Powder - ura g gg YP ml Difco gbecton Dickinson g g SC - ura GCY1 YPR1 SC - ura ml ml SC - ura ml OD nm YP ml SC - ura ml ml ml OD nm g g mlmm KCl mm phml

3 Saccharomyces cerevisiae g Vibrogen- Zellumuhle g g μm Bio-Rad Profinia ml IMAC Bio-Rad ml GST Bio- Rad Profinia Profinia μlμl μl Bio-Rad TGX V ORIOLE Bio-Rad SDS-PAGE ma PVDF HA Open Biosystems Invitrogen Western Breeze ml ml mm phml M ml M ml mm NAD ml ml U nm S. cerevisiae GCY1 YPR1 S. cerevisiae S. cerevisiae GCY1 YPR1 Costenoble et al. S. cerevisiae ORF BG Gelperin et al. BG GAL1 Gelperin et al. GCY1 YPR1 GCY1 GCYYPR1 YPR ph ph Cheng et al.hallborn et al.linlink et al.stein et al. Wong et al. GCY YPR GCY YPR His GCY1 YPR1 BG ORF His HA Protein A ZZ C GCY YPRBio-Rad

4 His Ni Profinia A B GCY1 C YPR1 A BC D nm Ni mlnm Profinia IMAC nm AB C D A D B GCY1 GCY D C YPR1 GCY1 GCY SDS-PAGE GCY1 KDa His KDa KDa D HA GCY YPR D GCY1 Profinia SDS MA B C D Profinia

5 Saccharomyces cerevisiae GCY1 YPR1 Profinia A GCY1 B YPR1 MA B C D Profinia GCY1 YPR1 A GCY1 B YPR1 D His GCY YPR Ni His GCY YPR GCY

6 Cellulomonas sp. G Enterobacter aerogenes G A Cellulomonas sp. GB Enterbacter aerogenes G YPR GST YPR1 pegh ORF N S GSTHis YPR1 GST GST-YPR GST-YPR A Cellulomonas sp. G B Enterobactor aerogenes G Oliveira et al.dominguez et al. Saccharomyces cerevisiae Kluyvermyces lactis S. cerevisiae N C Cheng H, Jiang N, Shen A, and Feng YMolecular cloning and functional expression of D-arabitol dehydrogenase gene from Gluconobacter oxydans in Escherichia coli. FEMS Microbiol. Lett. Costenoble R, Valadi H, Gustafsson L, Niklasson C, and Franzen CJ Microaerobic glycerol formation in Saccharomyces cerevisiae. Yeast

7 Saccharomyces cerevisiae de Figueroa LIC and Lucca MEProdction of polyols by omsmotolerant yeasts. Food Microbiol. Protocols Dominguez KBH, Toth IV, et al.sequential injection kinetic flow assay for monitoring glycerol in a sugar fermentation process by Saccharomyces cerevisiae. Appl. Biochem. Biotechnol. Dujon B et al.genome evolution in yeasts. Nature Gelperin DM, White MA, et al. Biochemical and genetic analysis of the yeast proteome with a movable ORF collection. Genes Developm. Hallborn J, Walfridsson M, Penttila M, Keranen S, and Hahn- Hagerdal B A Short-chain dehydrogenase gene from Pichia stipites having D-arabitoldehydrogenase activity. Yeast Iwaki T, Kurono S, Yokose Y, Kubota K, Tamai Y, and Watanabe Y Cloning of glycerol- -phosphate dehydrogenase gene ZrGPD1 and ZrGPD2 and glycerol dehydrogenase genes ZrGCY1 and ZrGCY2 from the salt-tolerant yeast Zygosaccharomyces rouxii. Yeast Lin ECCAn inducible D-arabitol dehydrogenase from Aerobacter aerogenes. J.Biol.Chem. Link T, Lohaus G, Heiser I, Mendgen K, Hahn M, and Voegele RTCharacterization of a novel NADP + dependent D-arabitol dehydrogenase from the plant pathogen Uromyces fabae. Biochem. J. Minamihara T and Suzuki MProperties of sorbitol dehydrogenase from Pseudomonas sp. KS-E and comparison with other sorbitiol dehydrogenases. J. Ferment. Bioeng. Oliveira HM, Segundo MA, et al.kinetic enzymatic determination of glycerol in wine and beer using a sequential injection system with spectrophotometric detection. J. Agric. Food Chem. Onishi HOsmophillic yeast. Adv. Food Res. Stein R, Gross W, and Schnarrenberger CCharacterization of a xylitol dehydrogenase and a D-arabitol dehydrogenase from the thermo- and acidophilic red alga Galdieria sulphuraria. Planta Tang XM, Kayingo G, and Prior BA Functional analysis of the Zygosaccharomyces rouxii Fps1p homologue. Yeast van Zyl PJ and Prior BAWater relations of polyol accumulation by Zygosaccharomyces rouxii in continuous culture. Appl. Microbiol. Biotechnol. van Zyl PJ, Kilian SG, and Prior PATheroleofan active transport mechanism in glycerol accumulation during osmoregulation by Zygosaccharomyces rouxii. Appl. Microbiol. Biotechnol. Watanabe Y, Nagayama K, and Tamai YExpession of glycerol -phosphate dehydrogenase gene CvGPD1 in salt-tolerant yeast Candida versatilis in stimulated by high concentrations of NaCl. Yeast Watanabe Y, Takechi Y, Nagayama K, and Tamai Y Overexpression of Saccharomyces cerevisiae mannitol dehydrogenase gene (YEL w) in glycerol synthesisdeficient S. cerevisiae mutant. Enzyme Microbial. Technol. Watanabe Y, Tsuchimoto S, and Tamai YHeterologous expression of Zygosaccharomyces rouxii glycerol phosphate dehydrogenase gene ZrGPD1 and glycerol dehydrogenase gene ZrGCY1 in Saccharomyces cerevisiae. FEMS Yeast Res. Wong B, Murray JS, Castellanos M, and Croen KD D-arabitol metabolism in Candida albicans: Studies of the biosynthetic pathway and the gene that encodes NAD - dependent D-arabitol dehydrogenase. J. Bacteriol. Food Function

Introduction ur company has just started service to cut out sugar chains from protein and supply them to users by utilizing the handling technology of

Standard PA-Sugar Chain Catalogue Masuda Chemical Industries Co., LTD. http://www.mc-ind.co.jp Introduction ur company has just started service to cut out sugar chains from protein and supply them to users