Identification and characterization of a putative endolysin encoded by episomal phage phiSM101 of Clostridium perfringens

• Published in: Applied microbiology and biotechnology 2011-06, Vol.90 (6), p.1973-1979
• Main Authors: Nariya, Hirofumi, Miyata, Shigeru, Tamai, Eiji, Sekiya, Hiroshi, Maki, Jun, Okabe, Akinobu
• Format: Article
• Language: English
• Subjects: Affinity chromatography; Analysis; Antibiotics; Antimicrobial agents; Applied Genetics and Molecular Biotechnology; Bacteria; Bacteriolysis; Bacteriophages - enzymology; Bacteriophages - isolation & purification; Biological and medical sciences; Biological control; Biotechnology; Chromatography; Chromatography, Affinity - methods; Cloning; Cloning, Molecular; Clostridium perfringens; Clostridium perfringens - virology; Dogs; E coli; Endopeptidases - genetics; Endopeptidases - isolation & purification; Endopeptidases - metabolism; Enzymes; Escherichia coli; Escherichia coli - genetics; Ethylenediaminetetraacetic acid; Fundamental and applied biological sciences. Psychology; Gene Expression; Genes; Genetic engineering; Genomes; Genomics; Glycoside Hydrolases - genetics; Glycoside Hydrolases - isolation & purification; Glycoside Hydrolases - metabolism; Gram-positive bacteria; Health aspects; Histidine; Life Sciences; Lysozyme; Microbial Genetics and Genomics; Microbiology; Mutation; Nickel; Pests; Recombinant Proteins - genetics; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Streptococcus infections; Studies; Surveys; Toxins; Turbidity
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-011-3253-z

Clostridium perfringens produces potent toxins and histolytic enzymes, causing various diseases including life-threatening fulminant diseases in humans and other animals. Aiming at utilizing a phage endolysin as a therapeutic alternative to antibiotics, we surveyed the genome and bacteriophage sequences of C. perfringens . A phiSM101 muramidase gene ( psm ) revealed by this study can be assumed to encode an N -acetylmuramidase, since the N-terminal catalytic domain deduced from the gene shows high homology of those of N -acetylmuramidases. The psm gene is characteristic in that it is present in phiSM101, an episomal phage of enterotoxigenic C. perfringens type A strain, SM101, and also in that homologous genes are present in the genomes of all five C. perfringens toxin types. The psm gene was cloned and expressed in Escherichia coli as a protein histidine-tagged at the N-terminus (Psm-his). Psm-his was purified to homogeneity by nickel-charged immobilized metal affinity chromatography and anion-exchange chromatography. The purified enzyme lysed cells of all C. perfringens toxin types but not other clostridial species tested, as was shown by a turbidity reduction assay. These results indicate the Psm-his is useful as a cell-wall lytic enzyme and also suggest that it is potentially useful for biocontrol of this organism.