Effect of Bacillus subtilis spo0A mutation on cell wall lytic enzymes and extracellular proteases, and prevention of cell lysis

• Published in: Journal of bioscience and bioengineering Vol. 103; no. 1; pp. 13 - 21
• Main Authors: Kodama, Takeko, Endo, Keiji, Ara, Katsutoshi, Ozaki, Katsuya, Kakeshita, Hiroshi, Yamane, Kunio, Sekiguchi, Junichi
• Format: Journal Article
• Language: English
• Published: Amsterdarm Elsevier B.V 2007; Elsevier Science; Elsevier Limited
• Subjects: ALFA AMILASA; ALPHA AMYLASE; autolysins; BACILLUS SUBTILIS; Bacterial Proteins - genetics; Bacteriolysis - physiology; Biological and medical sciences; Biotechnology; Cell Membrane - enzymology; Cell Membrane - genetics; CELL WALLS; ENZIMAS; ENZYME; ENZYMES; Escherichia coli - cytology; Escherichia coli - physiology; extracellular proteases; Fundamental and applied biological sciences. Psychology; GENE; GENES; Genetic Enhancement - methods; MUTANT; MUTANTES; MUTANTS; Mutation; N-Acetylmuramoyl-L-alanine Amidase - metabolism; PARED CELULAR; PAROI CELLULAIRE; Peptide Hydrolases - genetics; Peptide Hydrolases - metabolism; PROTEASAS; PROTEASE; PROTEASES; Protein Engineering - methods; spo0A; Transcription Factors - genetics; spo0A; extracellular proteases; Bacillus subtilis; autolysins; Enzyme; Metalloendopeptidases; Bacillaceae; Autolysin; Extracellular; Cell wall; Prevention; Peptidases; Bacillales; Lysis; Bacteria; Hydrolases; Mutation
• ISSN: 1389-1723; 1347-4421
• DOI: 10.1263/jbb.103.13

The Bacillus subtilis spo0A mutant is an adequate host for extracellular protein production ( e.g., α-amylase). However the mutant was prone to cell lysis. SDS–PAGE and zymography of cell wall lytic proteins indicated that the spo0A mutant contained high amounts of two major autolysins (LytC [CwlB] and LytD [CwlG]) and two minor cell wall lytic enzymes (LytE [CwlF] and LytF [CwlE]). On the other hand, the expression of eight extracellular protease genes was very poor or absent in the spo0A mutant. An eight-extracellular-protease-deficient mutant (Dpr8 strain) was constructed and the strain also exhibited cell lysis. The autolysins from the spo0A mutant were degraded by the supernatant of the wild type but not degraded by that of the Dpr8 mutant. These results suggest that the extensive cell lysis of the spo0A mutant was partially caused by the stability of autolysins via the decrease of the extracellular proteases. The introduction of a major autolysin and/or SigD mutations into the spo0A mutant was effective for preventing cell lysis.