Structure of Cruzipain/Cruzain Inhibitors Isolated from Bauhinia bauhinioides Seeds

Abstract

The saline extract of Bauhinia bauhinioides dry seeds was shown to inhibit cruzipain, a cysteine proteinase from Trypanosoma cruzi. The inhibitory activity was assigned to a protein with 164 amino acid residues and molecular mass of 18 034 Da that was purified by chromatography on DEAE-Sephadex, trypsin-Sepharose (removal of trypsin inhibitors), Mono Q and a reversed-phase C4 column. The primary structure is homologous to other plant Kunitz-type inhibitors, but it lacks cysteine residues and therefore the disulfide bridges. No methionine residue was identified by amino acid sequencing. The inhibition of cruzipain fits into a slow-tight binding mechanism with a low dissociation constant (Ki 1.2 nM). The studied Bauhinia protein also inhibits cruzain (Ki 0.3 nM), a C-terminally truncated recombinant species of cruzipain. Cathepsin L, a cysteine proteinase with high homology to cruzipain, is also inhibited (Ki 0.22 nM), but not cathepsin B, papain, bromelain or ficin.

Bauhinia bauhinioides cruzipain inhibitor reduces endothelial proliferation and induces an increase of the intracellular Ca2+ concentration

“…Furthermore, the antiproliferative effect involves rise in cytosolic Ca 2+ concentration and changes in membrane potential in response to BbCI stimulation. BbCI is a potent inhibitor of cruzipain from T. cruzi and also of the highly homologue mammalian proteinase cathepsin L [6,34]. Cruzipain is in many aspects similar to the human tissue kallikrein, e.g.…”

“…BbCI also shows inhibitory properties against some other serine proteinases: the porcine pancreatic elastase, the human neutrophil elastase and cathepsin G [2,30]. But interestingly, BbCI does not demonstrate a significant inhibition of the serine proteinase plasma kallikrein or of papain and cathepsin B both cysteine proteinases closely related with cruzipain [8].…”

“…Proteinase inhibitors from Bauhinia bauhinioides are known to effect physiological processes like blood coagulation, inflammation or cancer [29,30,38]. B. bauhinioides cruzipain inhibitor (BbCI) is a serine and cysteine proteinase inhibitor, which is isolated from the seeds of B. bauhinioides [8]. BbCI is an 18-kDa protein, an inhibitor of cruzipain-the main cysteine proteinase of Trypanosoma cruzi as well as its recombinant form cruzain and cathepsin L-a mammalian proteinase which is highly homologue to cruzipain [6,34].…”

“…B. bauhinioides cruzipain inhibitor (BbCI) is a serine and cysteine proteinase inhibitor, which is isolated from the seeds of B. bauhinioides [8]. BbCI is an 18-kDa protein, an inhibitor of cruzipain-the main cysteine proteinase of Trypanosoma cruzi as well as its recombinant form cruzain and cathepsin L-a mammalian proteinase which is highly homologue to cruzipain [6,34]. BbCI also shows inhibitory properties against some other serine proteinases: the porcine pancreatic elastase, the human neutrophil elastase and cathepsin G [2,30].…”

Immunomodulatory properties of cystatins

“…It has been shown recently that chagasin inhibits cruzipain [29 -30]. Surprisingly, BbCl is a cruzipain inhibitor isolated from Bauhinia bouhinoides seeds without any related structure with cystatins [31]. 1504 B. Vray, S. Hartmann and J. Hoebeke Immunomodulatory properties of cystatins Clearly, cystatins and cystatin-like molecules constitute a large and growing field of interest for biochemists, immunologists and parasitologists.…”