Fig. 7. CYK-3 and DOA4 specifically cleave ubiquitin from linear protein fusions. Ubiquitin (UB) and the ubiquitin-like proteins CeSUMO, NEDD-8 and a ubiquitin-like protein encoded by cosmid H06I04 were fused to a truncated version of ß-Gal (ß-Gal) and used as substrates. The fusion proteins were expressed in bacteria together with either CYK-3 or yeast DOA4. Cleavage activity was assessed by probing with an anti-ß-Gal-antibody on a western blot. Both CYK-3 and DOA4 specifically cleave ubiquitin from ß-Gal. In contrast, none of the ubiquitin-like proteins is cleaved from the ß-Gal substrate in the presence of CYK-3 or DOA4.