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Genetic analysis of the Bacillus licheniformis degSU operon and the impact of regulatory mutations on protease production

Paper ID Volume ID Publish Year Pages File Format Full-Text
23685 43463 2012 9 PDF Available
Title
Genetic analysis of the Bacillus licheniformis degSU operon and the impact of regulatory mutations on protease production
Abstract

Disruption experiments targeted at the Bacillus licheniformis degSU operon and GFP-reporter analysis provided evidence for promoter activity immediately upstream of degU. pMutin mediated concomitant introduction of the degU32 allele – known to cause hypersecretion in Bacillus subtilis – resulted in a marked increase in protease activity. Application of 5-fluorouracil based counterselection through establishment of a phosphoribosyltransferase deficient Δupp strain eventually facilitated the marker-free introduction of degU32 leading to further protease enhancement achieving levels as for hypersecreting wild strains in which degU was overexpressed. Surprisingly, deletion of rapG – known to interfere with DegU DNA-binding in B. subtilis – did not enhance protease production neither in the wild type nor in the degU32 strain. The combination of degU32 and Δupp counterselection in the type strain is not only equally effective as in hypersecreting wild strains with respect to protease production but furthermore facilitates genetic strain improvement aiming at biological containment and effectiveness of biotechnological processes.

► Protease hypersecretion. ► Marker-free mutagenesis. ► Counterselection. ► Degradative enzymes deregulation. ► Stationary phase heterogeneity.

Keywords
Hypersecretion; Uracil-phosphoribosyltransferase; Extracellular enzymes; Marker-free mutagenesis
First Page Preview
Genetic analysis of the Bacillus licheniformis degSU operon and the impact of regulatory mutations on protease production
Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 159, Issues 1–2, 31 May 2012, Pages 12–20
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering