Immunological study on integrated PilQ and disulphide loop region of PilA against acute Pseudomonas aeruginosa infection: In silico analysis and in vitro production

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• 作者：Alireza Salimi Chirani¹ ; ^{Alireza.SalimiChirani@sbmu.ac.ir" class="auth_mail" title="E-mail the corresponding author} ; Robabeh Majidzadeh² ; Hossein Dabiri¹ ; Javad Rezaei³ ; Ali Esmaili¹ ; Yasamin Abdanan Kord⁴ ; Narges Khabazzadeh Tehrani⁵ ; Negin Attaran⁴
• 关键词：Pseudomonas aeruginosa ; In silico analysis ; Type IV pili ; PilQ ; Vaccine ; Immunogen
• 刊名：Journal of Acute Disease
• 出版年：2016
• 出版时间：1 March 2016
• 年：2016
• 卷：5
• 期：2
• 页码：131-142
• 全文大小：2331 K

文摘

Nowadays, Pseudomonas aeruginosa (P. aeruginosa), the highly regarded opportunistic pathogen, is the leading cause of morbidity and mortality worldwide. The P. aeruginosa type IV pili (T4P) as a multiple functional surface organelle in the development of acute P. aeruginosa infections have been well documented. Today, in silico analysis is a quick, and cost-effective tool for vaccine development.

Methods

In present study, several turns' motifs along with the chimeric protein were predicted. Based on the hydropathy analysis, numerous antibody-accessible hydrophilic regions were characterized in the chimeric protein. A synthetic chimeric gene, encoding integrated PilQ and disulphide loop region of PilA, was designed. Modeling was done to predict the 3D structure of protein. The model was validated by using Ramachandran plot statistics and by ProSA server. Identification of B-cell and T-cell corresponding epitopes was done by using appropriate servers.

Results

The closer 3D model to the native form of the chimeric protein was achieved. Validation results showed that 95.1% residues were in favor region and 3.6% of amino acid residues were in the allowed region. The B-cell epitope mappings showed that almost all the epitopes had irregular enriched structures. The major histocompatibility complex binding sequence prediction identified several human major histocompatibility complex class I and II restricted T-cell epitopes. The integrated PilQ and PilA disulphide loop encoding regions in the frame of pET28a(+) vector were expressed and purified efficiently.

Conclusions

We expect that the two recognized antigenic determinants from our chimeric protein, “AYHKGNWSGYGKDGNIGIKDEDGMNCGPIAGSCTFPTTGTSKSPSPFVDLGAKDATSG” and “GPIAGSCTFPTTGTSKSPSP”, can be able to evoke strong both humoral and cell-mediated immune responses in mouse models.