Proteasome inhibitors have been described as an important target for
cancer therapy due to their potential to regulate the
ubiquitin-
proteasome system in the degradation pathway of cellular
proteins. Here, we reported the effects of a Bowman-Birk-type
protease inhibitor, the
Black-eyed pea Trypsin/Chymotrypsin Inhibitor (BTCI), on
proteasome 20S in MCF-7
breast cancer cells and on catalytic activity of the purified
20S proteasome from horse erythrocytes, as well as the structural analysis of the BTCI-20S
proteasome complex. In vitro experiments and confocal microscopy showed that BTCI readily crosses the membrane of the
breast cancer cells and co-localizes with the
proteasome in cytoplasm and mainly in nucleus. Indeed, as indicated by dynamic light scattering, BTCI and
20S proteasome form a stable complex at temperatures up to 55°C and at neutral and alkaline pHs. In complexed form, BTCI strongly inhibits the proteolytic
chymotrypsin-, trypsin- and
caspase-like activities of
20S proteasome, indicated by inhibition constants of 10(-7) M magnitude order. Besides other mechanisms, this feature can be associated with previously reported
cytostatic and cytotoxic effects of BTCI in MCF-7
breast cancer cells by means of apoptosis.