Prediction of superantigen active sites and clonal expression of staphylococcal enterotoxin-like W.
10.3760/cma.j.cn112338-20220822-00725
- Author:
Yu Hua YANG
1
;
Xin KU
2
;
Ya Nan GONG
1
;
Fan Liang MENG
1
;
Dong bo BU
3
;
Ya Hui GUO
4
;
Xiao Yue WEI
1
;
Li Jin LONG
5
;
Jia Ming FAN
1
;
Mao Jun ZHANG
1
;
Jian Zhong ZHANG
1
;
Xiao Mei YAN
1
Author Information
1. State Key Laboratory of Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China.
2. Key Lab of Intelligent Information Processing, Institute of Computing Technology, Chinese Academy of Sciences, Beijing 100190, China University of Chinese Academy of Sciences, Beijing 100049, China.
3. Key Lab of Intelligent Information Processing, Institute of Computing Technology, Chinese Academy of Sciences, Beijing 100190, China University of Chinese Academy of Sciences, Beijing 100049, China Big Data Academy, Zhongke, Zhengzhou 450046, China.
4. State Key Laboratory of Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China Baotou Medical College, Inner Mongolia University of Science and Technology, Baotou 014040, China.
5. State Key Laboratory of Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China China Medical University, Shenyang 110122, China.
- Publication Type:Journal Article
- MeSH:
Humans;
Enterotoxins/genetics*;
Superantigens/genetics*;
Catalytic Domain;
Selenoprotein W/metabolism*;
Receptors, Antigen, T-Cell
- From:Chinese Journal of Epidemiology
2023;44(4):629-635
- CountryChina
- Language:Chinese
-
Abstract:
Objective: The docking and superantigen activity sites of staphylococcal enterotoxin-like W (SElW) and T cell receptor (TCR) were predicted, and its SElW was cloned, expressed and purified. Methods: AlphaFold was used to predict the 3D structure of SElW protein monomers, and the protein models were evaluated with the help of the SAVES online server from ERRAT, Ramachandran plot, and Verify_3D. The ZDOCK server simulates the docking conformation of SElW and TCR, and the amino acid sequences of SElW and other serotype enterotoxins were aligned. The primers were designed to amplify selw, and the fragment was recombined into the pMD18-T vector and sequenced. Then recombinant plasmid pMD18-T was digested with BamHⅠand Hind Ⅲ. The target fragment was recombined into the expression plasmid pET-28a(+). After identification of the recombinant plasmid, the protein expression was induced by isopropyl-beta-D- thiogalactopyranoside. The SElW expressed in the supernatant was purified by affinity chromatography and quantified by the BCA method. Results: The predicted three-dimensional structure showed that the SElW protein was composed of two domains, the amino-terminal and the carboxy-terminal. The amino-terminal domain was composed of 3 α-helices and 6 β-sheets, and the carboxy-terminal domain included 2 α-helices and 7 antiparallel β-sheets composition. The overall quality factor score of the SElW protein model was 98.08, with 93.24% of the amino acids having a Verify_3D score ≥0.2 and no amino acids located in disallowed regions. The docking conformation with the highest score (1 521.328) was selected as the analysis object, and the 19 hydrogen bonds between the corresponding amino acid residues of SElW and TCR were analyzed by PyMOL. Combined with sequence alignment and the published data, this study predicted and found five important superantigen active sites, namely Y18, N19, W55, C88, and C98. The highly purified soluble recombinant protein SElW was obtained with cloning, expression, and protein purification. Conclusions: The study found five superantigen active sites in SElW protein that need special attention and successfully constructed and expressed the SElW protein, which laid the foundation for further exploration of the immune recognition mechanism of SElW.
