Characterization of a tissue kallikrein inhibitor isolated from Bauhinia bauhinioidesseeds: inhibition of the hydrolysis of kininogen related substrates

Trypsin inhibitors were purified from a saline extract of Bauhinia bauhinioidesseeds by ion-exchange column chromatography on DEAE-Sephadex, gel filtration on Superose 12 column, Mono Q ion-exchange chromatography or, alternatively, by affinity chromatography on trypsin-Sepharose. Both B. bauhinioidesisolated inhibitors, BbTI-Iand BbTI-II, inhibit trypsin being the dissociation constant 0.6 and 0.36 nM, respectively. BbTI-IIonly inhibits porcine pancreatic kallikrein hydrolysis of H-Pro-Phe-Arg-AMC (Ki2.0 nM); the bradykinin-containing sequence LGMISLMKRPPGFSPFRSSRI–NH2and the two kininogen related flanking quenched substrates Abz-MISLMKRP-EDDnp (Ki2.0 nM) and Abz-FRSSRQ-EDDnp (Ki2.5 nM).