fulltext.study @t Gmail

Enhancing the recombinant protein expression of halohydrin dehalogenase HheA in Escherichia coli by applying a codon optimization strategy

Paper ID Volume ID Publish Year Pages File Format Full-Text
17288 42657 2011 7 PDF Available
Title
Enhancing the recombinant protein expression of halohydrin dehalogenase HheA in Escherichia coli by applying a codon optimization strategy
Abstract

Halohydrin dehalogenases are attractive biocatalysts in producing a series of important chiral building blocks. Recombinant expression of halohydrin dehalogenase from Arthrobacter sp. AD2 (HheA) in Escherichia coli using T7 promoter-based pGEF(+) system revealed much lower expression level than that of the well-studied halohydrin dehalogenase from Agrobacterium radiobacter AD1 (HheC). In this study, we changed the codon usage in the 5′-end of hheA gene to improve the expression yield of HheA. Our results showed that the expression of HheA could be largely improved by the replacement of G-rich +2 codon (adjacent to the start codon) with less G-containing codons. The expression of one of the resulting mutants HheA-D1 (replaced +2 codon GTG with CCA) was about 4-fold higher and purified yields about 8-fold greater than that of the wild-type HheA. Moreover, the expression level of the resulting HheA variants correlated well with the minimal folding free energy (ΔG) of the mRNA secondary structure surrounding the 5′-end region of the genes. These findings suggested that the G-rich +2 codon of hheA gene might be the main suppressive factor for limiting the recombinant expression of HheA and that +2 codon optimization strategy could be used as a general tool in modulating recombinant protein production in E. coli.

• 5′-end codon optimization strategy is applied to improve the expression yield of HheA. • HheA expression is largely improved by G-rich +2 codon optimization. • 5′-end rare codon does not greatly affect HheA expression. • High HheA expression relates with poor stability of mRNA secondary structure at 5′ encoding region. • High expression level facilitates the purification of HheA.

Keywords
Halohydrin dehalogenase; Recombinant expression; +2 Codon optimization; pGEF(+) expression vector; Purification
First Page Preview
Enhancing the recombinant protein expression of halohydrin dehalogenase HheA in Escherichia coli by applying a codon optimization strategy
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
Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 49, Issue 4, 10 September 2011, Pages 395–401
Authors
, , , ,
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