Human bleomycin hydrolase binds ribosomal proteins

Bleomycin hydrolase (BH) is a cysteine proteinase that inactivates the anti cancer drug bleomycin, Yeast BH forms a homohexameric structure that resemb les a 20S proteasome and binds to single-stranded RNA and DNA. We now demon strate that human BH (hBH) interacts and colocalizes with ribosomal protein s. Using a yeast two-hybrid system, we found hBH bound to human homologues of rat ribosomal proteins L11 and L29. The N-terminus of hBH (amino acids 1 4-175), which contains a catalytic Cys(93), was critical for the binding to L11 in the two-hybrid environment. hBH precipitated S-35-labeled L11 and L 29 in vitro, and hBH colocalized with L11 and L29 as determined by immunofl uorescence. In addition to cytosolic bleomycin hydrolase, we found abundant bleomycin hydrolase activity associated with the ribosomal subcellular fra ction by differential centrifugation. hBH was also detected by Western immu noblotting in a high-speed particulate fraction, where the majority of L11 and L29 were found. Ln vitro experiments showed recombinant hBH binds to Ch inese hamster ovary cell microsomes. Thus, our data strongly suggest that h BH exists as both a free cytosolic and ribosome-associated protein.