Antigenicity of the orthologous proteins in the Brucella OMVs

From the 30 clusters of the 4 Brucella species, the prediction of antigenicity was performed in silico. The analysis focused on the proteins with superficial subcellular location in the outer membrane identified through the alignment of orthologous proteins to obtain a consensus sequence. The antigenicity of the proteins with subcellular location of outer membrane found in the OMVs was predicted by alignments to orthologous proteins to obtain a consensus sequence using UniProt UGENE V.1.30. The antigenicity of the consensus protein was analyzed with VaxiJen V.2.0 (http://www.ddg-pharmfac.net/vaxijen/VaxiJen/VaxiJen.html) using the default threshold value (Doytchinova and Flower 2007). In addition, B and T cell epitope predictions were performed for each protein using the BCPREDS server (http://ailab.ist.psu.edu/bcpred/predict.html), and the predictions were performed with a specificity of 80% and an epitope length of 20 amino acids (El-Manzalawy et al. 2008). In addition, MHCpred V.2.0 (http://www.ddg-pharmfac.net/mhcpred/MHCPred/) was used to predict the T cell epitopes based on binding affinities to the MHC-I and MHC-II molecules (Guan et al. 2003). The server was adjusted to predict epitopes with a binding affinity greater than 15 for DRB1*0101, the most common allele in the human population (Vishnu et al. 2017). The B and T cell epitope density in a given protein was calculated by dividing the number of predicted epitopes by the length of the protein. The cumulative score was calculated by adding the score obtained from the VaxiJen server and the B and T cell epitope density values (Hisham and Ashhab 2018).