Structural Basis for Cyclosporin Isoform-Specific Inhibition of Cyclophilins from Toxoplasma gondii

Abstract

Cyclosporin (CsA) has antiparasite activity against the human pathogen Toxoplasma gondii. A possible mechanism of action involves CsA binding to T. gondii cyclophilins, although much remains to be understood. Herein, we characterize the functional and structural properties of a conserved (TgCyp23) and a more divergent (TgCyp18.4) cyclophilin isoform from T. gondii. While TgCyp23 is a highly active cis–trans-prolyl isomerase (PPIase) and binds CsA with nanomolar affinity, TgCyp18.4 shows low PPIase activity and is significantly less sensitive to CsA inhibition. The crystal structure of the TgCyp23:CsA complex was solved at the atomic resolution showing the molecular details of CsA recognition by the protein. Computational and structural studies revealed relevant differences at the CsA-binding site between TgCyp18.4 and TgCyp23, suggesting that the two cyclophilins might have distinct functions in the parasite. These studies highlight the extensive diversification of TgCyps and pave the way for antiparasite interventions based on selective targeting of cyclophilins.

Insights Into Peptidyl-Prolyl Cis-Trans Isomerases From Clinically Important Protozoans: From Structure to Potential Biotechnological Applications

“…Moreover, among the recombinant CyPs from protist parasites, TvCyP19 and TvCyP19.9 from T. vaginalis exhibited the lowest PPIase activity. Their activity is lower than that of hCyPA [26,27,130], which could be attributed to differing substrate affinities. These data suggest that these TvCyPs might exhibit different activities on other substrates (Table 11).…”

“…Several CyPs, such as PfCyP19 and PfCyP22 from P. falciparum, LmCyP19 from L. major and TgCyP23 from T. gondii, demonstrated high levels of activity, comparable to those of hCyPA (Kcat/Km = 4.9 × 10 6 M -1 s -1 ) [116,130]. Moreover, among the recombinant CyPs from protist parasites, TvCyP19 and TvCyP19.9 from T. vaginalis exhibited the lowest PPIase activity.…”

“…Most CyPs from T. vaginalis retain the aa residues essential for catalytic activity within the highly conserved CLD identified in TvCyP19 (referred to as TvCyP1 by Hsu et al, 2014) [26]: H 62 , R 63 , F 68 , M 69 , Q 71 , G 80 , A 109 , N 110 , A 111 , Q 119 , F 121 , W 129 , L 130 , and H 134 . Exceptions to this pattern were observed in TvCyP21, TvCyP37, and TvCyP44 (Supplementary Figure S2).…”

“…The antiparasitic activity of CsA is more strongly associated with calcineurin inhibition than with PPIase inhibition [151]. To fully understand the mechanism by which CsA inhibits infection, it is necessary to identify the parasite CsA receptor [130]. An important part of parasite rPPIase studies is identifying whether known inhibitors (CsA, FK506, and rapamycin) or new inhibitors inhibit a wide variety of biological processes in which PPIases are involved.…”

“…Variations in affinities between PPIases and their inhibitors, such as TgCyP18.4 and TgCyP23, have been attributed to changes in crucial binding site residues. These alterations influence the affinities of these PPIases for CsA [130].…”