Structure of Cruzipain/Cruzain Inhibitors Isolated from Bauhinia bauhinioides Seeds

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 DEAESephadex, trypsinSepharose (removal of trypsin inhibitors), Mono Q and a reversed phase C[4] column. The primary structure is homologous to other plant Kunitztype 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 slowtight binding mechanism with a low dissociation constant (K 1.2 nM). The studied Bauhinia protein also inhibits cruzain (K 0.3 nM), a Cterminally truncated recombinant species of cruzipain. Cathepsin L, a cysteine proteinase with high homology to cruzipain, is also inhibited (K 0.22 nM), but not cathepsin B, papain, bromelain or ficin.