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Search Results (3)

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Keywords = uridine diphosphate dependent glucosyltransferases (UGTs)

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12 pages, 2934 KiB  
Article
Enhanced Heterologous Production of Glycosyltransferase UGT76G1 by Co-Expression of Endogenous prpD and malK in Escherichia coli and Its Transglycosylation Application in Production of Rebaudioside
by Wenju Shu, Hongchen Zheng, Xiaoping Fu, Jie Zhen, Ming Tan, Jianyong Xu, Xingya Zhao, Shibin Yang, Hui Song and Yanhe Ma
Int. J. Mol. Sci. 2020, 21(16), 5752; https://doi.org/10.3390/ijms21165752 - 11 Aug 2020
Cited by 28 | Viewed by 4497
Abstract
Steviol glycosides (SGs) with zero calories and high-intensity sweetness are the best substitutes of sugar for the human diet. Uridine diphosphate dependent glycosyltransferase (UGT) UGT76G1, as a key enzyme for the biosynthesis of SGs with a low heterologous expression level, hinders its application. [...] Read more.
Steviol glycosides (SGs) with zero calories and high-intensity sweetness are the best substitutes of sugar for the human diet. Uridine diphosphate dependent glycosyltransferase (UGT) UGT76G1, as a key enzyme for the biosynthesis of SGs with a low heterologous expression level, hinders its application. In this study, a suitable fusion partner, Smt3, was found to enhance the soluble expression of UGT76G1 by 60%. Additionally, a novel strategy to improve the expression of Smt3-UGT76G1 was performed, which co-expressed endogenous genes prpD and malK in Escherichia coli. Notably, this is the first report of constructing an efficient E. coli expression system by regulating prpD and malK expression, which remarkably improved the expression of Smt3-UGT76G1 by 200% as a consequence. Using the high-expression strain E. coli BL21 (DE3) M/P-3-S32U produced 1.97 g/L of Smt3-UGT76G1 with a yield rate of 61.6 mg/L/h by fed-batch fermentation in a 10 L fermenter. The final yield of rebadioside A (Reb A) and rebadioside M (Reb M) reached 4.8 g/L and 1.8 g/L, respectively, when catalyzed by Smt3-UGT76G1 in the practical UDP-glucose regeneration transformation system in vitro. This study not only carried out low-cost biotransformation of SGs but also provided a novel strategy for improving expression of heterologous proteins in E. coli. Full article
(This article belongs to the Special Issue Molecular Enzymology: Advances and Applications)
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24 pages, 14592 KiB  
Article
Characterization of the UDP-glycosyltransferase UGT72 Family in Poplar and Identification of Genes Involved in the Glycosylation of Monolignols
by Nathanael Speeckaert, Nassirou Mahamadou Adamou, Hadjara Amadou Hassane, Fabien Baldacci-Cresp, Adeline Mol, Geert Goeminne, Wout Boerjan, Pierre Duez, Simon Hawkins, Godfrey Neutelings, Thomas Hoffmann, Wilfried Schwab, Mondher El Jaziri, Marc Behr and Marie Baucher
Int. J. Mol. Sci. 2020, 21(14), 5018; https://doi.org/10.3390/ijms21145018 - 16 Jul 2020
Cited by 35 | Viewed by 4904
Abstract
Monolignols are the building blocks for lignin polymerization in the apoplastic domain. Monolignol biosynthesis, transport, storage, glycosylation, and deglycosylation are the main biological processes partaking in their homeostasis. In Arabidopsis thaliana, members of the uridine diphosphate-dependent glucosyltransferases UGT72E and UGT72B subfamilies have [...] Read more.
Monolignols are the building blocks for lignin polymerization in the apoplastic domain. Monolignol biosynthesis, transport, storage, glycosylation, and deglycosylation are the main biological processes partaking in their homeostasis. In Arabidopsis thaliana, members of the uridine diphosphate-dependent glucosyltransferases UGT72E and UGT72B subfamilies have been demonstrated to glycosylate monolignols. Here, the poplar UGT72 family, which is clustered into four groups, was characterized: Group 1 UGT72AZ1 and UGT72AZ2, homologs of Arabidopsis UGT72E1-3, as well as group 4 UGT72B37 and UGT72B39, homologs of Arabidopsis UGT72B1-3, glycosylate monolignols. In addition, promoter-GUS analyses indicated that poplar UGT72 members are expressed within vascular tissues. At the subcellular level, poplar UGT72s belonging to group 1 and group 4 were found to be associated with the nucleus and the endoplasmic reticulum. However, UGT72A2, belonging to group 2, was localized in bodies associated with chloroplasts, as well as possibly in chloroplasts. These results show a partial conservation of substrate recognition between Arabidopsis and poplar homologs, as well as divergent functions between different groups of the UGT72 family, for which the substrates remain unknown. Full article
(This article belongs to the Special Issue Carbohydrate-Active Enzymes: Structure, Activity and Reaction Products 2020)
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11 pages, 1393 KiB  
Article
Biocatalytic Synthesis of Calycosin-7-O-β-D-Glucoside with Uridine Diphosphate–Glucose Regeneration System
by Yumei Hu, Jian Min, Yingying Qu, Xiao Zhang, Juankun Zhang, Xuejing Yu and Longhai Dai
Catalysts 2020, 10(2), 258; https://doi.org/10.3390/catal10020258 - 20 Feb 2020
Cited by 8 | Viewed by 3885
Abstract
Calycosin-7-O-β-D-glucoside (Cy7G) is one of the principal components of Radix astragali. This isoflavonoid glucoside is regarded as an indicator to assess the quality of R. astragali and exhibits diverse pharmacological activities. In this study, uridine diphosphate-dependent glucosyltransferase (UGT) UGT88E18 was [...] Read more.
Calycosin-7-O-β-D-glucoside (Cy7G) is one of the principal components of Radix astragali. This isoflavonoid glucoside is regarded as an indicator to assess the quality of R. astragali and exhibits diverse pharmacological activities. In this study, uridine diphosphate-dependent glucosyltransferase (UGT) UGT88E18 was isolated from Glycine max and expressed in Escherichia coli. Recombinant UGT88E18 could selectively and effectively glucosylate the C7 hydroxyl group of calycosin to synthesize Cy7G. A one-pot reaction by coupling UGT88E18 to sucrose synthase (SuSy) from G. max was developed. The UGT88E18–SuSy cascade reaction could recycle the costly uridine diphosphate glucose (UDPG) from cheap sucrose and catalytic amounts of uridine diphosphate (UDP). The important factors for UGT88E18–SuSy cascade reaction, including UGT88E18/SuSy ratios, different temperatures, and pH values, different concentrations of dimethyl sulfoxide (DMSO), UDP, sucrose, and calycosin, were optimized. We produced 10.5 g L−1 Cy7G in the optimal reaction conditions by the stepwise addition of calycosin. The molar conversion of calycosin was 97.5%, with a space–time yield of 747 mg L−1 h−1 and a UDPG recycle of 78 times. The present study provides a new avenue for the efficient and cost-effective semisynthesis of Cy7G and other valuable isoflavonoid glucosides by UGT–SuSy cascade reaction. Full article
(This article belongs to the Special Issue Biocatalytic Process Optimization)
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