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Cascade synthesis of uridine-5'-diphosphate glucuronic acid by coupling multiple whole cells expressing hyperthermophilic enzymes.


ABSTRACT: BACKGROUND:Enzymatic glycan synthesis has leapt forward in recent years and a number of glucuronosyltransferase (EC 2.4.1.17) have been identified and prepared, which provides a guide to an efficient approach to prepare glycans containing glucuronic acid (GlcA) residues. The uridine 5'-diphosphate (UDP) activated form, UDP-GlcA, is the monosaccharide donor for these glucuronidation reactions. RESULTS:To produce UDP-GlcA in a cost-effective way, an efficient three-step cascade route was developed using whole cells expressing hyperthermophilic enzymes to afford UDP-GlcA from starch. By coupling a coenzyme regeneration system with an appropriate expression level with UDP-glucose 6-dehydrogenase in a single strain, the cells were able to meet NAD+ requirements. Without addition of exogenous NAD+, the reaction produced 1.3 g L-1 UDP-GlcA, representing 100% and 46% conversion of UDP-Glc and UTP respectively. Finally, an anion exchange chromatography purification method was developed. UDP-GlcA was successfully obtained from the cascade system. The yield of UDP-GlcA during purification was about 92.0%. CONCLUSIONS:This work built a de novo hyperthermophilic biosynthetic cascade into E. coli host cells, with the cells able to meet NAD+ cofactor requirements and act as microbial factories for UDP-GlcA synthesis, which opens a door to large-scale production of cheaper UDP-GlcA.

SUBMITTER: Meng DH 

PROVIDER: S-EPMC6604206 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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Cascade synthesis of uridine-5'-diphosphate glucuronic acid by coupling multiple whole cells expressing hyperthermophilic enzymes.

Meng Dan-Hua DH   Du Ran-Ran RR   Chen Lu-Zhou LZ   Li Meng-Ting MT   Liu Fei F   Hou Jin J   Shi Yi-Kang YK   Wang Feng-Shan FS   Sheng Ju-Zheng JZ  

Microbial cell factories 20190701 1


<h4>Background</h4>Enzymatic glycan synthesis has leapt forward in recent years and a number of glucuronosyltransferase (EC 2.4.1.17) have been identified and prepared, which provides a guide to an efficient approach to prepare glycans containing glucuronic acid (GlcA) residues. The uridine 5'-diphosphate (UDP) activated form, UDP-GlcA, is the monosaccharide donor for these glucuronidation reactions.<h4>Results</h4>To produce UDP-GlcA in a cost-effective way, an efficient three-step cascade rout  ...[more]

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