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Enhanced extracellular expression of gene-optimized Thermobifida fusca cutinase in Escherichia coli by optimization of induction strategy

Paper ID Volume ID Publish Year Pages File Format Full-Text
34349 45019 2015 8 PDF Available
Title
Enhanced extracellular expression of gene-optimized Thermobifida fusca cutinase in Escherichia coli by optimization of induction strategy
Abstract

•IPTG and lactose co-induced the expression of T. fusca cutinase.•An extracellular cutinase activity of 2258.5 U mL−1 (5.1 g L−1) was obtained.•This cutinase activity represents the highest yield reported to date.•This study showed the potential industrial production of cutinase.

Our previous study demonstrated that when Thermobifida fusca cutinase was expressed in Escherichia coli without mediation of a signal peptide, it could release to the culture medium via the enhanced membrane permeability, which was based on its limited phospholipid hydrolysis activity. The goal of the present work was to achieve highly efficient extracellular production of the recombinant signal peptide-free cutinase in E. coli. The codons of the T. fusca cutinase gene were optimized for expression in E. coli, and a recombinant expression system was constructed using this optimized gene. After that, the induction strategy was optimized using high-cell-density cultivation in a 3-L fermentor. Results showed that the optimal induction condition was at a dry cell weight (DCW) of 13 g L−1, and an IPTG/lactose combination induction strategy is proposed, in which IPTG is added once in a final concentration of 25.0 μM, and lactose is fed at a rate of 0.5 g L−1 h−1. In this condition, an extracellular cutinase activity of 2258.5 U mL−1 (5.1 g L−1) was achieved, which represented the highest cutinase production ever reported, and demonstrated the potential of this system for the industrial production of cutinase.

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Keywords
Cutinase; Escherichia coli; Extracellular expression; Codon optimization; Induction strategy
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Enhanced extracellular expression of gene-optimized Thermobifida fusca cutinase in Escherichia coli by optimization of induction strategy
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Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 50, Issue 7, July 2015, Pages 1039–1046
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us