Abstract
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.