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

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• 作者：Claudia Borgmeier^a ; Johannes Bongaerts^b ; Friedhelm Meinhardt^a ; ^{meinhar@uni-muenster.de}
• 关键词：Hypersecretion ; Uracil-phosphoribosyltransferase ; Extracellular enzymes ; Marker-free mutagenesis
• 刊名：Journal of Biotechnology
• 出版年：2012
• 期刊代码：53_01681656
• 类别：imm
• 出版时间：31 May, 2012
• 卷：159
• 期：1-2
• 页码：12-20
• 文件大小：1074 K

摘要

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 螖upp 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 螖upp 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.