Production of protease with Bacillus licheniformis mutants insensitive to repression of exoenzyme biosynthesis

Two mutant strains of Bacillus licheniformis insensitive to catabolite repression were selected by classical mutagenesis in connection with the development of a fed-batch procedure for protease production. B. licheniformis 4a produced up to 20 U (Anson-Units) subtilisin Carlsberg/ml in fed-batch experiments in the presence of up to 1.5 m glycerol, but was inhibited by excess ammonium. Formation of spores, excretion of α-amylase and the biosynthesis of citrate synthase and isocitrate dehydrogenase were likewise not repressed by glycerol. The strain was characterized by unusually low activity of the α-oxoglutarate dehydrogenase complex and increased biosynthesis of polyglutamic acid in the presence and excretion of α-oxoglutarate in the absence of ammonium, respectively. The results are discussed in view of a possible connection between the defect in the α-oxoglutarate dehydrogenase complex and insensitivity to catabolite repression. The second strain B. licheniformis 114 was able to synthesize 11.5 U protease/ml independently of the glycerol and ammonium concentration in the medium.