Toxoplasma gondii aspartic protease 5: structural basis of substrate binding and inhibition mechanism

Abstract

Toxoplasma gondii, a worldwide prevalent parasite is responsible for causing toxoplasmosis in almost all warm-blooded animals, including humans. Golgi-resident T. gondii aspartic protease 5 (TgASP5) plays an essential role in the maturation and export of the effector proteins those modulate the host immune system to establish a successful infection. Hence, inhibiting this enzyme can be a possible therapeutic strategy against toxoplasmosis. This is the first report of the detailed structural investigations of the TgASP5 mature enzyme using molecular modeling and an all-atom simulation approach which provide in-depth knowledge of the active site architecture of TgASP5. The analysis of the binding mode of the TEXEL (Toxoplasma EXport Element) substrate to TgASP5 highlighted the importance of the active site residues. Ser505, Ala776 and Tyr689 in the S2 binding pocket are responsible for the specificity towards Arg at the P2 position of TEXEL substrate. The molecular basis of inhibition by the only known inhibitor RRL_Statine has been identified, and our results show that it blocks the active site by forming a hydrogen bond with a catalytic aspartate. Besides that, known aspartic protease inhibitors were screened against TgASP5 using docking, MD simulations and MM-PBSA binding energy calculations. The top-ranked inhibitors (SC6, ZY1, QBH) showed higher binding energy than RRL_Statine. Understanding the structural basis of substrate recognition and the binding mode of these inhibitors will help to develop potent mechanistic inhibitors against TgASP5. This study will also provide insights into the structural features of pepsin-like aspartic proteases from other apicomplexan parasites for developing antiparasitic agents.

Communicated by Ramaswamy H. Sarma

Keywords:

• TgASP5
• Toxoplasma
• aspartic protease
• TEXEL substrate
• apicomplexan parasite
• molecular dynamics
• RRL_Statine inhibitor

Acknowledgements

We would like to thank Spacetime High-Performance Computing (HPC) facility at Indian Institute of Technology Bombay (IIT Bombay), Mumbai, India for the computational time. SC thanks IIT Bombay for PhD fellowship. AD thanks to UGC for PhD fellowship. PK thanks IIT Bombay for financial support.

Data availability statement

Data associated with the study are available from the authors upon request.

Disclosure statement

Authors declare no competing financial interest.

Additional information

Funding

The author(s) reported there is no funding associated with the work featured in this article.