Journal
BIOCHEMICAL ENGINEERING JOURNAL
Volume 68, Issue -, Pages 1-6Publisher
ELSEVIER
DOI: 10.1016/j.bej.2012.07.003
Keywords
Co-expression; BioBrick vector; Cellulase; Enzyme production; Biodegradation; Enzyme activity
Funding
- National Natural Science Foundation of China [21176215/21176102]
- Outstanding Young Scholar of Zhejiang Province [R4110092]
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The enzymes used for biomass decomposition comprise a major cost in the production of biofuels from lignocellulosic feedstocks. Engineering of key enzymes de novo in heterologous hosts provides one strategy for the rational improvement of enzyme cocktails. Until recently, Escherichia coli has remained the most commonly used host for recombinant protein expression. Nevertheless, to our knowledge, there are few reports describing the co-expression of biomass degrading enzymes in E. coli. In this study, bicistronic and dual-promoter constructs based on pET30a were built for the co-expression of an endo-beta-xylanase gene (xyn) and a beta-glucosidase gene (bgl) from Trichoderma reesei QM 9414 in E. coli. The internal ribosome binding site used in the bicistronic constructs was originally found in pET30a. In the dual-promoter constructs described here, a pET30a-derived BioBrick base vector was built for the standard assembly of two targeted genes. Compared with monocistronic constructs, the crude enzyme expressed from a bicistronic construct (xyn located upstream of bgl) and a dual-promoter construct (xyn located upstream of bgl) offered the comparable activity of two recombinant proteins. Our results indicated that the common commercial vectors, such as pET30a, could be modified and optimized for a particular co-expression strategy in E. coli. (C) 2012 Elsevier B.V. All rights reserved.
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