Identification and characterization of a putative endolysin encoded by episomal phage phiSM101 of Clostridium perfringens
Identification and characterization of a putative endolysin encoded by episomal phage phiSM101 of Clostridium perfringens
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DOI：
10.1007/s00253-011-3253-z
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发表时间：
2011-06-01
影响因子：
5
通讯作者：
Okabe, Akinobu
中科院分区：
文献类型：
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作者：
Nariya, Hirofumi;Miyata, Shigeru;Okabe, Akinobu
产气荚膜梭菌产生强效毒素和溶组织酶，导致人类和其他动物的各种疾病，包括危及生命的暴发性疾病。为了利用噬菌体内溶素作为抗生素的治疗替代品，我们调查了C.产气荚膜杆菌phiSM 101胞壁酶基因（psm）与N-乙酰胞壁酶的N-末端催化结构域具有很高的同源性，因此推测该基因编码N-乙酰胞壁酶。psm基因的特征在于它存在于phiSM 101中，phiSM 101是产肠球菌的附加体噬菌体。A型产气荚膜梭菌菌株SM 101，并且还在于同源基因存在于所有五种C.产气荚膜杆菌毒素类型。克隆psm基因，并在大肠杆菌中表达为N-末端组氨酸标记的蛋白质（Psm-his）。Psm-his经镍固定化金属亲和层析和阴离子交换层析纯化至均一。纯化的酶裂解所有的C.产气荚膜杆菌毒素类型，而不是测试的其他梭菌物种，如浊度降低测定所示。这些结果表明，Psm-his是有用的细胞壁裂解酶，也表明它是潜在的有用的生物防治该生物体。
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.