Amino acid residue Y196E substitution and C-terminal peptide synergistically alleviate the toxicity of Clostridium perfringens epsilon toxin.
Epsilon toxin (ETX) is produced by Clostridium perfringens type B and D strains, and is the causative agent of a lethal enterotoxemia in livestock animals and possibly in humans. However, many details of ETX structure and activity are not known. Therefore, it is important to clarify the relationship between ETX structure and activity. To explore the effect and mechanism of ETX amino acid residue Y196E substitution and C-terminal peptide on toxicity, four recombinant proteins, rETX (without 13 N-terminal peptides and 23 C-terminal peptides), rETX-C (rETX with 23 C-terminal peptides), rETX(Y196E) (rETX with an amino acid residue substitution at Y196) and rETX(Y196E)-C (rETX-C with a Y196E mutation), were constructed in this study. Both the amino acid residue Y196E substitution and the C-terminal peptide reduce ETX toxicity to a similar extent, and the two factors synergistically alleviate ETX toxicity. In addition, we demonstrated that the C-terminal peptides and Y196E amino acid mutation reduce the toxin toxicity in two different pathways: the C-terminal peptides inhibit the binding activity of toxins to target cells, and the Y196E amino acid mutation slightly inhibits the pore-forming or heptamer-forming process. Interaction between the two factors was not observed in pore-forming or binding assays but toxicity assays, which demonstrated that the relationship between domains of the toxin is more complicated than previously appreciated. However, the exact mechanism of synergistic action is not yet clarified.
State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, 100071, Beijing, PR China.
Wenwu
W
Yao
State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, 100071, Beijing, PR China.
Lin
L
Kang
State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, 100071, Beijing, PR China.
Shan
S
Gao
State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, 100071, Beijing, PR China.
Xiangjin
X
Zhuang
State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, 100071, Beijing, PR China.
Tao
T
Zhang
State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, 100071, Beijing, PR China.
Hao
H
Yang
State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, 100071, Beijing, PR China.
Bin
B
Ji
State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, 100071, Beijing, PR China. Electronic address: xinww@hotmail.com.
Wenwen
W
Xin
State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, 100071, Beijing, PR China. Electronic address: wjl0801@vip.163.com.
Jinglin
J
Wang
D019943:Amino Acid Substitution; D000818:Animals; D001427:Bacterial Toxins / Q000737:chemistry* / Q000633:toxicity; D015153:Blotting, Western; D002942:Circular Dichroism; D004285:Dogs; D005260:Female; D061985:Madin Darby Canine Kidney Cells; D008807:Mice, Inbred BALB C; D016297:Mutagenesis, Site-Directed; D018675:Toxicity Tests
D001427:Bacterial Toxins; C039811:Clostridium perfringens epsilon-toxin
Clostridium perfringens; Enterotoxemia; Epsilon toxin (ETX); Substitution; Toxicity