Developing Multi-epitope Antigen Construct from Immunodominant Proteins for Serological Diagnosis of Chlamydia trachomatis: An In Silico Approach

Background: Chlamydiasis is a widespread bacterial infection in the world. Serological tests are expensive, and in addition, intrinsic antigens can cause cross-reactions and make the diagnosis process difficult. Multi-epitope protein antigens are novel and potential diagnostic markers that have the capability of more accurate and cheaper diagnosis. Therefore, in this study, the main goal is to design a new protein vaccine, including multiple epitopes of B cells with dominant immunity from three proteins named MOMP, ompA and Pgp3D from C. trachomatis Methods: The amino acid sequences were obtained from the UniProt database. The areas with the highest antigenicity were identified using the EMBOSS server. Linear B cell epitopes were determined using BCPRED, ABCpred, and Bepipred servers. Epitopes with the highest antigenicity were connected using the EAAAK linker. Results: Two epitopes from MOMP, two from ompA, and one from Pgp3D were selected. These epitopes were connected to each other with the EAAAK linker. Three residues (0.592), 16 residues (0.76), 36 residues (0.578), and 37 residues (0.734) were obtained from the prediction of the spatial structure of the B cell multiple epitopes designed with ElliPro. Model 1 of RaptorX was selected as the best structure. In this model, the ERRAT quality, ProSA-web z-score, and Verify3D were 83.1169, - 5.17 and 84.62% with PASS score, respectively. Moreover, the Ramachandran plot showed that 86.093% of the amino acid residues were located in the favored region. To achieve the highest level of protein expression, the designed multi-epitope reverse-translated with the Genscript server and was cloned in E. coli. The highest level of expression was achieved, and a CAI score of 0.91 was reported. The gene GC content was 51.98%, and the contribution of low-frequency codons was 0%. Conclusion: The results confirmed that the designed construct could identify C. trachomatis with high sensitivity and specificity in serum samples of patients with chlamydiasis. However, further experimental studies are needed for final confirmation.