Characterization and Pharmacological Properties of a Novel Multifunctional Kunitz Inhibitor from Erythrina velutina Seeds

• DOI: 10.1371/journal.pone.0063571
• Publication Date: 2013-05-29T03:58:21Z
• Authors: Richele J. A. Mac...; Norberto K. V. Mo...; Ludovico Migliolo; Osmar N. Silva; Michele F. S. Pinto; Adeliana S. Oliveira; Octávio L. Franco; Sumika Kiyota; Marcelo P. Bemquerer; Adriana F. Uchoa; Ana H. A. Morais; Elizeu A. Santos
• Keywords: Trypsin inhibitor

Inhibitors of peptidases isolated from leguminous seeds have been studied for their pharmacological properties. The present study focused on purification, biochemical characterization and anti-inflammatory anticoagulant evaluation a novel Kunitz trypsin inhibitor Erythrina velutina (EvTI). Trypsin inhibitors were purified by ammonium sulfate (30-60%), fractionation followed Trypsin-Sepharose affinity chromatography reversed-phase high performance liquid chromatography. showed molecular mass 19,210.48 Da. Furthermore, second isoform with 19,228.16 Da was also observed. that highest specificity enhanced recovery yield named EvTI (P2) selected further analysis. peptide fragments, generated pepsin digestion, analyzed MALDI-ToF-ToF spectrometry, allowing partial primary structure elucidation. exhibited inhibitory activity against IC50 2.2×10(-8) mol.L(-1) constant inhibition (Ki) 1.0×10(-8) mol.L(-1), non-competitive mechanism. In addition to inhibit the trypsin, inhibited factor Xa neutrophil elastase, but do not thrombin, chymotrypsin or peptidase 3. investigated its anti-coagulant Firstly, no cytotoxic effect human peripheral blood cells. Nevertheless, able prolong clotting time in dose-dependent manner using vitro vivo models. Due properties, two sepsis models here challenged. leukocyte migration specifically acted inhibiting TNF-α release stimulating IFN-α IL-12 synthesis. data presented clearly contribute better understanding use as bioactive agent capable interfering coagulation inflammation.