Annotating the ‘hypothetical’ in hypothetical proteins: In-silico analysis of uncharacterised proteins for the Apicomplexan parasite, Neospora caninum

Neospora caninum is a parasite of veterinary and economic importance, affecting beef and dairy cattle industries globally. While this species has been recognised as a serious cause of disease in cattle and dogs for over 30 years, treatment and control options are still not available. Furthermore, whilst vaccination was identified as the most economic control strategy, vaccine discovery programs require new leads to investigate as vaccines.The current lack of gene annotation available for N. caninum, especially compared to the closely related model organism, Toxoplasma gondii, considerably hinders vaccine related research. Moreover, due to the high degree of similarity between the two organisms, a significant amount of gene annotation available for N. caninum stems from sequence homology between the species. However, there is a plethora of literature identifying conserved virulence factors between members of the Apicomplexa, which suggests that key players are contributing to successful parasite invasion, motility, and host cell attachment.In this study, bioinformatic approaches classified 125 uncharacterised proteins within the N. caninum genome, as transmembrane proteins with signal peptide sequences. Functional annotation assigned enriched gene ontologies for cell-adhesion, ATP binding, protein serine/threonine phosphatase complex, immune system process, antigen binding, and proteolysis. Additionally, 32 of these proteins were also identified as adhesins, or having adhesin-like properties, which were further characterised through the discovery of domains and gene ontology, to reveal their potential functional significance as virulence factors for N. caninum. This study identifies a new, small subset of proteins within N. caninum, that may be involved in host-cell interaction, parasite adhesion, and invasion, thereby implicating them as potential targets to exploit in the development of control options against the disease.