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

, a worldwide prevalent parasite is responsible for causing toxoplasmosis in almost all warm-blooded animals, including humans. Golgi-resident aspartic protease 5 (ASP5) 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 ASP5 mature enzyme using molecular modeling and an all-atom simulation approach which provide in-depth knowledge of the active site architecture of ASP5. The analysis of the binding mode of the TEXEL ( EXport Element) substrate to ASP5 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 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 ASP5 using docking, MD simulations and MM-PBSA binding energy calculations. The top-ranked inhibitors (SC6, ZY1, QBH) showed higher binding energy than RRL. Understanding the structural basis of substrate recognition and the binding mode of these inhibitors will help to develop potent mechanistic inhibitors against ASP5. 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.