Bioinformatics-based Design of a Multi-epitope Chimeric Vaccine Candidate Targeting Toxoplasma gondii

The development of an effective vaccine against Toxoplasma gondii (T. gondii) remains complex and challenging due to the biological complexity and antigenic diversity of this parasite and requires further study. Therefore, this study aimed to design a novel chimeric protein as a vaccine candidate containing four immunogenic antigens (GRA1, SAG1, ROP18, and MIC13).

Amino acid positions from GRA1 (81–180), SAG1 (201–300), ROP18 (321–420), and MIC13 (201–300) were selected and linked using A(EAAAK)A. The secondary and tertiary structures, antigenicity, allergenicity, physicochemical properties, codon optimization, and the secondary structure of the mRNA of the GSRM (GRA1–SAG1–ROP18–MIC13) protein were analyzed using various bioinformatics tools.

The GSRM protein had 442 amino acids with a Molecular Weight (MW) of 47,953.46 Da and a theoretical Isoelectric Point (pI) of 5.61. An aliphatic index of 80.11 was obtained for the construct. With an instability index of 43.07, the protein appears to exhibit moderate stability. The GRAVY of the protein was estimated to be -0.316, indicating that the protein is hydrophilic. The results also showed that this protein is antigenic and non-allergenic. Based on the evaluation of tertiary structures, GSRM was selected as the best structure for the vaccine candidate. Furthermore, analysis of the secondary structure of mRNA (ΔG = -387.70 kcal/mol) revealed the absence of stable hairpin structures at the 5' end, and therefore translation is possible.

In silico analyses indicate that GSRM possesses favorable immunological and physicochemical properties. Combining antigenic fragments from different life stages of T. gondii may enhance the effectiveness of the immune response.

The accuracy and precision of the in silico analyses should be confirmed by subsequent experimental studies in vitro and in vivo.