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Tackling codon usage bias for heterologous expression in Rhodobacter sphaeroides by supplementation of rare tRNAs

Paper ID Volume ID Publish Year Pages File Format Full-Text
16881 42620 2015 10 PDF Available
Title
Tackling codon usage bias for heterologous expression in Rhodobacter sphaeroides by supplementation of rare tRNAs
Abstract

•Eight rare tRNAs were selected and successfully expressed in Rhodobacter sphaeroides.•Supplied tRNAs remarkably improved expressions of two rare codon containing genes.•tRNA supplementation shows better performances in minimal medium than in rich medium.

The photosynthetic Rhodobacter species are promising alternative expression hosts in bioproduction and biorefinery due to their unique metabolic capacities. With prominent inner membrane areas and efficient endogenous translocation machineries, they are especially attractive for membrane protein expression. However, codon usage bias could be a limitation in the engineering of Rhodobacter species and has seldom been investigated. In this study, we tackled the codon bias of Rhodobacter by functionally expressing 8 rare tRNAs of Rhodobacter sphaeroides with a multi-copy vector. The impact of tRNA supplementation was evaluated through monitoring expression levels of two heterologous proteins with different phylogenetic origins, a membrane subunit of the riboflavin transporter, RibU, from Lactobacillus acidophilus La-14 and a decaheme cytochrome, MtrA, from Shewanella oneidensis. Our results showed that the performances were closely related to medium composition and rare codon percentages of raw DNA sequences. Provision of rare tRNAs has increased RibU production by 7.7-folds and 2.86-fold in minimal medium and rich medium, respectively, while MtrA levels were increased by 1-fold in minimal medium. The present study confirms the presence of codon bias in R. sphaeroides and offers a facile tool for improving heterologous expression of rare-codon containing genes. We anticipate that this tRNA supplementation system can be further extended to other species of Rhodobacter, and thus will facilitate the engineering of purple bacteria for interesting applications in microbial technology.

Keywords
Codon bias; Bacterial expression system; Heterologous expression level; Rhodobacter sphaeroides; tRNA level
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Tackling codon usage bias for heterologous expression in Rhodobacter sphaeroides by supplementation of rare tRNAs
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 72, May 2015, Pages 25–34
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