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The direct repeat sequence upstream of Bacillus chitinase genes is cis-acting elements that negatively regulate heterologous expression in E. coli

Paper ID Volume ID Publish Year Pages File Format Full-Text
17137 42646 2012 7 PDF Available
Title
The direct repeat sequence upstream of Bacillus chitinase genes is cis-acting elements that negatively regulate heterologous expression in E. coli
Abstract

To explore the influence of the direct repeat sequence (DRS) in Bacillus chitinase genes on heterogonous expression in Escherichia coli, we cloned and sequenced the entire open reading frame (ORF) and upstream sequences of the chitinase B (chiB) and chitinase MY75 (chiMY75) from Bacillus thuringiensis and Bacillus licheniformis. A pair of 8-bp DRS was found upstream of each chi gene. Chi ORFs with a series of truncated DRS were cloned and transformed into E. coli XL-Blue. The activity of the transformants without the DRS were significantly higher in chitinase assays than transformants containing the DRS. SDS-PAGE showed that part and full deletion of the DRS increased chi gene expression by approximately 1.7 and 3.8-fold, respectively. Northern blotting revealed deletion of the DRS regions increased chiB and chiMY75 mRNA expression. Specific binding of DNA-binding factors in the E. coli cell lyaste was observed to both the chiB and chiMY75 promoter regions and DRS elements. This is the first investigation to demonstrate that heterologous expression of Bacillus chi genes in E. coli is negatively regulated by their upstream DRS regions, which act as cis-acting elements.

Graphical abstractFigure optionsDownload full-size imageDownload as PowerPoint slideHighlights► A pair of 8-bp DRS was found upstream of two Bacillus chitinase genes. ► Part and full deletion of the DRS increased chi gene expression in Escherichia coli. ► DRS of Bacillus chitinase gene act as cis-acting elements in E. coli. ► Our results provide clues as to how E. coli governs expression of invading genes.

Keywords
Bacillus; Chitinase genes; Direct repeat sequence; Heterologous expression; Cis-acting elements
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The direct repeat sequence upstream of Bacillus chitinase genes is cis-acting elements that negatively regulate heterologous expression in E. coli
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 50, Issues 6–7, 10 May 2012, Pages 280–286
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