A target within the target: probing cruzain’s P1′ site to define structural determinants for the Chagas’ disease protease

Abstract

The series of vinyl-sulfone-based inhibitors examined in complex with cruzain was designed to probe recognition and binding potential of an aromatic-rich region of the enzyme. Analysis of the interactions formed shows that aromatic interactions play a less significant role, whereas the strength and importance of hydrogen bonding in the conformation adopted by the inhibitor upon binding to the enzyme was highlighted. A derivative of one inhibitor examined is currently under development as a therapeutic agent against Chagas' disease.

Toward the discovery of inhibitors of babesipain-1, a Babesia bigemina cysteine protease: in vitro evaluation, homology modeling and molecular docking studies

“…6a); this is an important feature for the enzyme's proteolytic activity, as the ''oxyanion hole'' stabilizes the tetrahedral adduct during the nucleophilic attack of the thiolate anion to the appropriate electron deficient carbonyl of the substrate [32]. Additionally, we observed the typical glycine-rich region, comprising mainly Gly65 and Gly66, that in other papain-like cysteine proteases was found to provide additional stability to the complex by forming a constellation of hydrogen bonds with the substrate [39].…”

Synthesis of macrocyclic trypanosomal cysteine protease inhibitors

“…Several cruzain-inhibitor complexes have been solved by X-ray crystallography, which displayed the active site Cys25 of cruzain covalently bound to the vinyl sulfone unit of the inhibitor. 16 With the aim to improve upon the lead compounds from previous studies and to develop an inhibitor with a broad spectrum of activity against a variety of parasitic hosts, we were interested in the design of conformationally constrained vinyl sulfones. Limiting conformational flexibility of the inhibitor or ligand is a well-established strategy to improve binding energies by decreasing the entropic barrier to binding of a particular conformation.…”

“…Examination of the crystal structures of cruzain with vinyl sulfones such as K11002 (1) reveal several highly conserved binding interactions. 16 These include hydrogen bonding between the side chains of Gln19, His159, and Trp177 with the sulfonyl oxygen atoms, a hydrogen bond between the P 1 nitrogen with the Asp158 peptide carbonyl, a hydrogen bond between the P 2 carbonyl and the Gly66 amide, and a hydrogen bond between the P 2 amide nitrogen with the Gly66 carbonyl ( Figure 1b). 16 The S 2 pocket is the primary recognition element for cruzain and all other enzymes in the papain class.…”

“…In addition, introduction of a linking unit between these two units was expected not to interfere with enzyme binding, since the P 1 homoPhe residue is highly solvent exposed in the available crystal structures. 16 We anticipated that 2 would not be chemically stable, due to the acid labile aminal moiety. This was addressed by replacing the aminal unit with an α-amino amide as shown in 3.…”

“…Our current hypothesis is that P 3 unit in 4 and 5 is sub-optimal compared to the P 3 units in K11002 and other dipeptidyl vinylsulfonyl inhibitors. 16 It is possible that the free amine in 4 or 5, which will be protonated under conditions of the enzyme assay, may negatively impact the binding of these inhibitors to the enzyme targets. It is also possible that the urea units of acyclic cysteine protease inhibitors (c.f., K11002) are much more important to inhibitor binding than initially assumed.…”