Identification of Proteins Responsible for High Activity of Cysteine Proteinase Inhibitor in the Blood of Nile Tilapia Oreochromis niloticus

Abstract

Cysteine proteinase inhibitors (CPIs) protect tissues and organs against cysteine proteinases in animal blood and have attracted much attention for use in food processing and medical sciences for humans and animals. Several CPI proteins, which include stefins, cystatins, kininogens, histidine-rich glycoproteins (HRG) and fetuins, have been identified and characterized in mammals. Fish blood also contains high CPI activity, but the identity of the major protein responsible for this activity has not been clarified. This study was conducted to screen CPI activity by examining papain inhibitory activity from various different tissues in Nile tilapia Oreochromis niloticus and to identify major proteins for the activity in the blood. CPI activity was highest in the serum among the tissues screened in this study (at least fourfold higher than in other tissues)(P < 0.05). Major proteins for CPI activity in serum were purified using a CNBr-activated sepharose 4B column, gel filtration and an ion exchange FPLC column. From these purifications, two proteins with strong CPI activity were isolated and partially sequenced. Based on their molecular weights and partial amino sequences, the two major proteins with CPI activity from the blood in this species were found to be fetuin B (60 kDa) and kininogen (54 kDa).