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Autolysis of lactic acid bacteria (LAB) plays a vital role in dairy processing. During cheese making, autolysis of LAB affects cheese flavor development through release of intracellular enzymes and restricts the proliferation of cells in yogurt fermentation and probiotics production. In order to explore the mechanism of autolysis, the gene for the autolytic enzymes of  L. bulgaricus , N-acetylmuramidase ( mur ), was cloned and sequenced (GenBank accession number: KF157911).  Mur  gene overexpression and gene knockout vectors were constructed based on pMG76e and pUC19 vectors. Recombinant plasmids were transformed into  L. bulgaricus  ljj-6 by electroporation, then three engineered strains with pMG76e-mur vector and fifteen engineered strains with pUC19-mur::EryBII were screened. The autolysis of the  mur  knockout strain was significantly lower and autolysis of the mur overexpressed strain was significantly higher compared with that of the wild type strain ljj-6. This result suggested that the  mur  gene played an important role in autolysis of  L. bulgaricus . On the other hand, autolytic activity in a low degree was still observed in the  mur  knockout strain, which implied that other enzymes but autolysin encoded by  mur  were also involved in autolysis of  L. bulgaricus .

Gene Knockout and Overexpression Analysis Revealed the Role of N-Acetylmuramidase in Autolysis of Lactobacillus delbrueckii subsp. bulgaricus Ljj-6