Amimoto Katsuhiko, Noro Taichi, Oishi Eiji, Shimizu Mitsugu
Kyoto Biken Laboratories, Inc., 24-16 Makishima-cho, Uji, Kyoto 611-0041, Japan.
Microbiology (Reading). 2007 Apr;153(Pt 4):1198-1206. doi: 10.1099/mic.0.2006/002287-0.
An unknown cytotoxin was identified in the culture supernatant of Clostridium perfringens type C. The cytotoxin, named TpeL, which was purified using mAb-based affinity chromatography, had a lethal activity of 62 minimum lethal dose (MLD) mg(-1) in mice and a cytotoxic activity of 6.2x10(5) cytotoxic units (CU) mg(-1) in Vero cells. The nucleotide sequence of TpeL was determined. The entire ORF had a length of 4953 bases, and the same nucleotide sequence was not recorded in the GenBank/EMBL/DDBJ databases. The molecular mass calculated from the deduced amino acid sequence was 191 kDa, and a signal peptide region was not found within the ORF. The deduced amino acid sequence exhibited 30-39 % homology to Clostridium difficile toxins A (TcdA) and B (TcdB), Clostridium sordellii lethal toxin (TcsL) and Clostridium novyi alpha-toxin (TcnA). The amino acid sequence of TpeL is shorter than these toxins, and the homologous region was located at the N-terminal site. Eighteen strains of C. perfringens types A, B and C were surveyed for the presence of the tpeL gene by PCR. The tpeL gene was detected in all type B (one strain) and C strains (five strains), but not in any type A strains (12 strains). TpeL was detected in culture filtrates of the five type C strains by dot-blot analysis, but not in the type B strain. It was concluded that TpeL is a novel toxin similar to the known large clostridial cytotoxins. Furthermore, the data indicated that TpeL is produced by many C. perfringens type C strains.
在产气荚膜梭菌C型的培养上清液中鉴定出一种未知的细胞毒素。该细胞毒素名为TpeL，通过基于单克隆抗体的亲和色谱法进行纯化，在小鼠体内具有62最小致死剂量（MLD）mg⁻¹的致死活性，在Vero细胞中具有6.2×10⁵细胞毒性单位（CU）mg⁻¹的细胞毒性活性。测定了TpeL的核苷酸序列。整个开放阅读框（ORF）长度为4953个碱基，在GenBank/EMBL/DDBJ数据库中未记录相同的核苷酸序列。根据推导的氨基酸序列计算出的分子量为191 kDa，在ORF内未发现信号肽区域。推导的氨基酸序列与艰难梭菌毒素A（TcdA）和B（TcdB）、索氏梭菌致死毒素（TcsL）和诺维氏梭菌α毒素（TcnA）具有30 - 39%的同源性。TpeL的氨基酸序列比这些毒素短，同源区域位于N端位点。通过PCR检测了18株A型、B型和C型产气荚膜梭菌中tpeL基因的存在情况。在所有B型（1株）和C型菌株（5株）中检测到了tpeL基因，但在任何A型菌株（12株）中均未检测到。通过斑点印迹分析在5株C型菌株的培养滤液中检测到了TpeL，但在B型菌株中未检测到。得出的结论是，TpeL是一种类似于已知大梭菌细胞毒素的新型毒素。此外，数据表明TpeL由许多产气荚膜梭菌C型菌株产生。
