• Journal of Ginseng Research
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• 제24권2호
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• pp.83-88
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• 2000

6년생 고려인삼근(Panax ginseng C.A. Meyer) 중의 Sucrose synthase, UDP-glucose pyrophosphorylase 및 ADP-glucose pyrophosphorylase의 활성을 생육 시기별로 조사한 결과, Sucrose synthase 와 ADP-glucose pyrophosphorylase는 뿌리저장활성 지표로서 adaptive enzyme의 특성을 나타내는 반면, UDP-glucose pyrophosphorylase는 maintenance enzyme으로서 존재하였다. 평균기온이 24。C 이상일 때 전분합성이 저하되고 중심부의 산소소비량이 급격히 증가되었다.

• KSBB Journal
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• 제21권5호
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• pp.366-370
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• 2006

UDP-glucose는 ${\beta}$-1,3-glucan 합성의 중요한 전구체로 이를 측정함으로써 세포 내의 glucan synthesis 대사의 활성도를 추정할 수 있는 중요한 지표가 됨을 본 연구 결과를 통하여 알 수 있었다. UDP-glucose는 세포 성장기에 다량 생산되다가 ${\beta}$-1,3-glucan 합성하는 시기에 일정한 농도가 되며 ${\beta}$-1,3-Glucan 합성 메카니즘에서 glucose를 운반하는 중요한 역할을 하는 것으로 나타났다. 2단 연속 발효조를 이용하여 세포 성장 발효조와 ${\beta}$-1,3-glucan 생산 발효조에서 UDP-glucose 변화를 관찰하여 ${\beta}$-1,3-glucan 생산시 농도가 높음을 관찰할 수 있었다. ${\beta}$-1,3-Glucan 생산 발효조의 pH를 5.5로 조절함으로써 UDP-glucose의 농도를 증가시킬 수 있을 뿐만 아니라 ${\beta}$-1,3-glucan의 생산 속도를 최적화할 수 있었다.

• KSBB Journal
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• 제23권6호
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• pp.474-478
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• 2008

효율적인 UDP-glucose regeneration system을 구축하기 위해서 재순환 시스템에 관여하는 4가지 효소 (UDP-glucose pyrophosphorylase, UDP-Kinase gene, UDP-galactose 4-epimerase, and $\beta$-1, 4-galactasyltrasnsferase)들을 E. coli AD202에서 발현 시켜 Disaccharide 합성 정도를 보았다. Disaccharide는 0.5 mM IPTG 농도에서 가장 높은 농도를 나타내었다. 대조구와 비교한 결과 LacNAc 농도는 1.34 mM로 10배 정도 정가하였고, lactose 농도는 0.39 mM로 대조구보다 2.6배 증가하였다. 총 disaccharide 농도는 1.73 mM 이며, 대조구 보다 6.5배 높은 생산성을 보였다. 본 논문은 결과는 metabolic flux regeneration으로 disaccharides 합성을 증가시킬 수 있다는 것을 보여주었다.

• BMB Reports
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• 제37권4호
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• pp.503-506
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• 2004

Glucose-1-phosphate uridylyltransferase from E. coli K12 was used to convert uridine-5'-triphosphate and glucose-1-phosphate to UDP-D-glucose. The conversion was efficient and completed within 5 minutes under the employed conditions. In addition, thymidine-5'-monophosphate kinase and acetate kinase were proven to be non-specific, converting udridine-5'-monophosphate to uridine-5'-triphosphate with 55% conversion after 6 h, which was much slower than the production of TTP under the same conditions (complete conversion within one hour). Since these two reactions could proceed under the same conditions, a one-pot synthesis of UDP-D-glucose with ATP regeneration was designed from easily available starting materials, and conversion up to 40% by HPLC peak integration was achieved given a reaction time of 4 h.

• Bulletin of the Korean Chemical Society
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• 제30권7호
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• pp.1547-1552
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• 2009

Sphingomonas chungbukensis DJ77 has the ability to produce large quantities of an extracellular polysaccharide that can be used as a gelling agent in the food and pharmaceutical industries. We identified, cloned and expressed the UDP-glucose dehydrogenase gene of S. chungbukensis DJ77, and characterized the resulting protein. The purified UDP-glucose dehydrogenase (UGDH), which catalyzes the reversible conversion of UDP-glucose to UDPglucuronic acid, formed a homodimer and the mass of the monomer was estimated to be 46 kDa. Kinetic analysis at the optimal pH of 8.5 indicated that the $K_m\;and\;V_{max}$ for UDP-glucose were 0.18 mM and 1.59 mM/min/mg, respectively. Inhibition assays showed that UDP-glucuronic acid strongly inhibits UGDH. Site-directed mutagenesis was performed on Gly9, Gly12 Thr127, Cys264, and Lys267. Substitutions of Cys264 with Ala and of Lys267 with Asp resulted in complete loss of enzymatic activity, suggesting that Cys264 and Lys267 are essential for the catalytic activity of UGDH.

• Journal of Microbiology and Biotechnology
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• 제19권7호
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• pp.709-712
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• 2009

Enzymatic glucosylation with glycosyltransferases can be used to regulate the water solubility of aglycone. The drawback of this process is the demand of UDP-glucose as a sugar donor. We made an in-frame fusion of the flavonoid O-glucosyltransferase (OsUGT-3) and sucrose synthase (AtSUS) genes. The resulting fusion protein, OsUGT3-AtSUS, was expressed in E. coli and purified. When sucrose and UDP were supplied, the fusion protein was able to convert quercetin into quercetin O-glucoside without the addition of UDP-glucose. In addition, UDP-glucose was recycled when sucrose was added to the reaction mixture. This fusion protein is useful for the enzymatic production of flavonoid O-glucosides.

• Journal of Microbiology and Biotechnology
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• 제12권2호
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• pp.217-221
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• 2002

dTDP-D-glucose 4,6-dehydratase (TDPDH) catalyzes the conversion of dTDP-D-glucose to dTDP-4-keto-6-deoxy-D-glucose, and requires $NAD^＋$ as a coenzyme for its catalytic activity. The dTDP-D-glucose 4,6-dehydratase from Streptomyces antibioticus $Tu{\ddot}99$ tightly binds $NAD^＋$ [19]. In order to determine the role of lysine-148 in the $NAD^＋$ binding, the lysine of the dTDP-D-glucose 4,6-dehydratase from Streptomyces antibioticus $Tu{\ddot}99$ was mutated to various amino acids by site-directed mutagenesis. The catalytic activity of the four mutated enzymes of TDPDH did not recover after addition of $NAD^＋$ . However, the activity of K159A, the mutated enzyme of UDP-D-glucose 4-epimerase (UDPE), recovered after the addition of $NAD^＋$ [15]. Although dTDP-glucose 4,6-dehydratase, and UDP-galactose (glucose) 4-epimerase are members of the short-chain dehydrogenase/reductase SDR family and the lysine-148 of TDPDH was highly conserved as in UDPE (Lys-159), the function of the lysine-148 of TDPDH was different from that of UDPE. The mutated enzymes showed that the lysine-148 of the dTDP-D-glucose 4,6-dehydratase played no role in the $NAD^＋$ binding. Accordingly, it is suggested that the lysine-148 of the dTDP-D-glucose 4,6-dehydratase is involved in the folding of TDPDH.

• Bulletin of the Korean Chemical Society
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• 제30권6호
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• pp.1360-1364
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• 2009

The bacterium Sphingomonas chungbukensis DJ77 produces the extracellular polysaccharide gellan in high yield. Gellan produced by this bacterium is widely used as a gelling agent, and the enzyme UDP-glucose pyrophosphorylase (UGP) is thought to play a key role in the gellan biosynthetic pathway. The UGP gene has been successfully cloned and over-expressed in E. coli. The expressed enzyme was purified with a molecular weight of approximately 32 kDa, as determined by a SDS-polyacrylamide gel, but the enzyme appears as ca. 63 kDa on a native gel, suggesting that the enzyme is present in a homodimer. Kinetic analysis of UDP-glucose for UGP indicates $K_m$ = 1.14 mM and $V_{max}$ = 10.09 mM/min/mg at pH 8.0, which was determined to be the optimal pH for UGP catalytic activity. Amino acid sequence alignment against other bacteria suggests that the UGP contains two conserved domains: An activator binding site and a glucose-1-phosphate binding site. Site-directed mutagenesis of Lys194, located within the glucose-1-phosphate binding site, indicates that substitution of the charge-reversible residue Asp for Lys194 dramatically impairs the UGP activity, supporting the hypothesis that Lys194 plays a critical role in the catalysis.

• 한국잠사곤충학회지
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• 제40권2호
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• pp.105-110
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• 1998

The baculovirus egt gene encodes an ecdysteroid UDP-glucosyltransferase(EGT) which catalyzes the transfer of glucose from UDP-glucose to the insect moltion hormone ecdysteroid resulting in a functionally inactive ecdysteroid. In baculovirus-infected insect larvae, EGT has been shown block molting and pupation. In this study, we compared the development of 4th and 5th instar silkworm, Bombyx mori, larvae injected with either wild-type bombyx mori nucleopolyhedrovirus (BmNPV) or a mutant BmNPV(BmEGTZ) in which the egt gene was disrupted by the insertion of a lacZ gene cassette. Larvae injected with BmEGTZ died roughly 12 h more rapidly compared to indentical larvae infected with BmNPV. In addition, BmEGTZ- infected larvae prematurely stopped feeding and gain less weight compared to BmNPV-infected larvae. In order to investigate why BmEGTZ-infected larvae died more rapidly than BmNPV-infected larvae, the array of hemolymph proteins in BmEGTZ-or BmNPV-infected larvae were analyzed by SDS-PAGE. The hemolymph of BmEGTZ-infected larvae showed virus-specific proteins, including polyhedrin, about 12 h earlier than the hemolymph of BmNPV-infected larvae

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2005년도 생물공학의 동향(XVII)
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• pp.674-679
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• 2005

With addition of uracil during fermentation, the production rate of $1,3-{\beta}-glucan(curdlan)$ was enhanced. Since uracil was used as precursor of UDP-Glucose, the UDP-glucose level was increased and further glucan synthesis rate was increased.