Comparative proteomics and structure-based approach to unravel the therapeutic drug target of Theileria species

Abstract

Theileriosis, caused by protozoan parasites of genus Theileria, primarily affects both domestic and wild ruminants. It can lead to significant economic losses in livestock farming due to decreased productivity and high mortality rates in susceptible animals, while treatment measures are not cost-effective. Since most of mechanisms of this disease remain unknown, this study investigates the differences in the mode of pathogenesis between transforming and non-transforming groups through an in silico comparative proteomics approach to recognize the key players involved in host cell transformation. Although the major biological processes and molecular functions are almost conserved between the two groups, PEST-motif containing secretory proteins of SfiI, SVSP, and Tash-AT gene families were identified as important candidates with the potential to transform infected host cells. Several members of PEST-motif containing proteins possess signal peptides, nuclear localization signals, and trans-membrane helices, further supporting their potential to transform host cells. Additionally, structural analysis helped in the identification of a parasitic protein (SfiIp) from SfiI family as a plausible drug target. Virtual screening revealed FDA-approved drugs (i.e. atogepant and rimegepant) as promising compounds, showing the highest affinity for SfiIp during molecular docking. Further studies, including molecular dynamics simulation, principal component analysis, and free energy landscape, suggested that these drug molecules exhibit the stable interaction with protein. Therefore, our research could facilitate the identification and targeting of novel drug candidates that may be further implemented to recognize effective therapeutics against Theileria infections.

Keywords: Comparative proteomics; Molecular dynamics simulations; SfiIp; Theileria; Transforming species.

Fig. 1

Schematic representation of the comparative proteome and structure-based analysis for identifying the drug targets and inhibitors of Theileria species.

Fig. 2

Phylogenetic relationship of eleven apicomplexan species wherein numbers indicate the Bootstrap values. P. falciparum branches out from the clade constituting Theileria and Babesia species with B. bovis being the last common ancestor of all four Theileria species.

Fig. 3

Biological process distribution of three Theileria species. The green, red, and purple circles correspond to Theileria annulata, Theileria orientalis, and Theileria equi. The major biological processes remain mostly conserved in all three species with majority of the proteins participating in various metabolic processes and gene expression. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Fig. 4

Cellular components distribution of proteomes of Theileria annulata (A), Theileria orientalis (B), and Theileria equi (C).

Fig. 5

Molecular function distribution of Theileria species. Theileria annulata, Theileria orientalis, and Theileria equi are represented by green, red, and purple circles, respectively. The proteins were predicted to primarily participate in catalytic activities and the binding of their cognate ligands across all three species. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Fig. 6

Distribution of (a) protein domains, (b) superfamilies, (c) sites and (d) repeats in proteomes of Theileria annulata (green), Theileria orientalis (pale blue), and Theileria equi (orange). PEST motif (IPR011695) is characteristic to only the transforming species and absent in other species (Theileria orientalis and Theileria equi). Notably, the DUF529 repeat (IPR007480) is unique to Theileria genus and present in the three species. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Fig. 7

(A) Modelled three-dimensional structure of SfiIp, which is a PEST motif-containing protein and encoded by a member of SfiI gene family in Theileria annulata. Alpha helices, beta sheets and coils are coloured in cyan, magenta and orange, respectively. (B) Ramachandran plot of SfiIp obtained through PROCHECK server. (C) Overall and (D) local model quality of refined SfiIp structure calculated from ProSA webserver. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Fig. 8

Interaction analysis of SfiIp with lead molecules. Docking of SfiIp was performed with FDA-approved drugs through AutoDock Vina, wherein protein is presented as white coloured surface view, while atogepant, and rimegepant are represented as yellow, and magenta coloured sticks, respectively. The interacting residues of protein with atogepant (A) and rimegepant (B) are shown as white sticks, whereas black dashed lines represent the polar contacts. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Fig. 9

MDS studies of apo SfiIp and its complexes with lead molecules. Comparative assessment of RMSD (A), Rg (B), RMSF (C), and SASA (D) of apo, atogepant and rimegepant bound SfiIp. Illustration of hydrogen bonds formed for SfiIp complexes (E) during the simulation period of 100 ns.

Fig. 10

Principal component analysis (PCA) plots constructed by eigenvector 1 vs eigenvector 2 of apo SfiIp (A) and its complexes with atogepant (B), and rimegepant (C). The combination of plots for apo (black), atogepant (red) and rimegepant (blue) bound SfiIp (D). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)