NDLI: Systematic comparison of co-expression of multiple recombinant thermophilic enzymes in Escherichia coli BL21(DE3)

Content Provider	Springer Nature Link
Author	Chen, Hui Huang, Rui Zhang, Y. H. Percival
Copyright Year	2017
Abstract	The precise control of multiple heterologous enzyme expression levels in one Escherichia coli strain is important for cascade biocatalysis, metabolic engineering, synthetic biology, natural product synthesis, and studies of complexed proteins. We systematically investigated the co-expression of up to four thermophilic enzymes (i.e., α-glucan phosphorylase (αGP), phosphoglucomutase (PGM), glucose 6-phosphate dehydrogenase (G6PDH), and 6-phosphogluconate dehydrogenase (6PGDH)) in E. coli BL21(DE3) by adding T7 promoter or T7 terminator of each gene for multiple genes in tandem, changing gene alignment, and comparing one or two plasmid systems. It was found that the addition of T7 terminator after each gene was useful to decrease the influence of the upstream gene. The co-expression of the four enzymes in E. coli BL21(DE3) was demonstrated to generate two NADPH molecules from one glucose unit of maltodextrin, where NADPH was oxidized to convert xylose to xylitol. The best four-gene co-expression system was based on two plasmids (pET and pACYC) which harbored two genes. As a result, apparent enzymatic activities of the four enzymes were regulated to be at similar levels and the overall four-enzyme activity was the highest based on the formation of xylitol. This study provides useful information for the precise control of multi-enzyme-coordinated expression in E. coli BL21(DE3).
Starting Page	4481
Ending Page	4493
Page Count	13
File Format	PDF
ISSN	01757598
Journal	Applied Microbiology and Biotechnology
Volume Number	101
Issue Number	11
e-ISSN	14320614
Language	English
Publisher	Springer Berlin Heidelberg
Publisher Date	2017-03-01
Publisher Place	Berlin, Heidelberg
Access Restriction	Subscribed
Subject Keyword	Co-expression Expression control Cascade biocatalysis E. coli BL21(DE3) T7 terminator Microbiology Microbial Genetics and Genomics Biotechnology
Content Type	Text
Resource Type	Article
Subject	Medicine Applied Microbiology and Biotechnology Biotechnology

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