PDBsum entry 4klb

Hydrolase/hydrolase inhibitor

PDB id: 4klb

Contents

Protein chains

215 a.a.

Ligands

1RV ×3

Waters ×109

PDB id:

4klb

Name:

Hydrolase/hydrolase inhibitor

Title:

Crystal structure of cruzain in complex with the non-covalent inhibitor nequimed176

Structure:

Cruzipain. Chain: a, b, c, d, e. Fragment: cruzain mature domain, unp residues 123-337. Synonym: cruzaine, major cysteine proteinase. Engineered: yes

Source:

Trypanosoma cruzi. Organism_taxid: 5693. Gene: cruzipain, genbank m84342.1. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

2.62Å R-factor: 0.192 R-free: 0.239

Authors:

W.B.Fernandes,C.A.Montanari,J.H.Mckerrow

Key ref:

H.J.Wiggers et al. (2013). Non-peptidic cruzain inhibitors with trypanocidal activity discovered by virtual screening and in vitro assay. Plos Negl Trop Dis, 7, e2370. PubMed id: 23991231 DOI: 10.1371/journal.pntd.0002370

Date:

07-May-13 Release date: 18-Sep-13

Protein chains

P25779  (CYSP_TRYCR) -  Cruzipain from Trypanosoma cruzi

Seq: 467 a.a.

Struc: 215 a.a.

Enzyme reactions

• Enzyme class: E.C.3.4.22.51 - cruzipain.

DOI no: 10.1371/journal.pntd.0002370

Plos Negl Trop Dis 7:e2370 (2013)

PubMed id: 23991231

Non-peptidic cruzain inhibitors with trypanocidal activity discovered by virtual screening and in vitro assay.

H.J.Wiggers, J.R.Rocha, W.B.Fernandes, R.Sesti-Costa, Z.A.Carneiro, J.Cheleski, A.B.da Silva, L.Juliano, M.H.Cezari, J.S.Silva, J.H.McKerrow, C.A.Montanari.

ABSTRACT

A multi-step cascade strategy using integrated ligand- and target-based virtual screening methods was developed to select a small number of compounds from the ZINC database to be evaluated for trypanocidal activity. Winnowing the database to 23 selected compounds, 12 non-covalent binding cruzain inhibitors with affinity values (K i) in the low micromolar range (3-60 µM) acting through a competitive inhibition mechanism were identified. This mechanism has been confirmed by determining the binding mode of the cruzain inhibitor Nequimed176 through X-ray crystallographic studies. Cruzain, a validated therapeutic target for new chemotherapy for Chagas disease, also shares high similarity with the mammalian homolog cathepsin L. Because increased activity of cathepsin L is related to invasive properties and has been linked to metastatic cancer cells, cruzain inhibitors from the same library were assayed against it. Affinity values were in a similar range (4-80 µM), yielding poor selectivity towards cruzain but raising the possibility of investigating such inhibitors for their effect on cell proliferation. In order to select the most promising enzyme inhibitors retaining trypanocidal activity for structure-activity relationship (SAR) studies, the most potent cruzain inhibitors were assayed against T. cruzi-infected cells. Two compounds were found to have trypanocidal activity. Using compound Nequimed42 as precursor, an SAR was established in which the 2-acetamidothiophene-3-carboxamide group was identified as essential for enzyme and parasite inhibition activities. The IC50 value for compound Nequimed42 acting against the trypomastigote form of the Tulahuen lacZ strain was found to be 10.6±0.1 µM, tenfold lower than that obtained for benznidazole, which was taken as positive control. In addition, by employing the strategy of molecular simplification, a smaller compound derived from Nequimed42 with a ligand efficiency (LE) of 0.33 kcal mol(-1) atom(-1) (compound Nequimed176) is highlighted as a novel non-peptidic, non-covalent cruzain inhibitor as a trypanocidal agent candidate for optimization.