Bleomycin hydrolase is a unique thiol aminopeptidase

Summary Bleomycin hydrolase, which hydrolyzes the car☐amide bond in the pyrimidoblamic acid moiety of the bleomycin molecule, also cleaved several p -nitroanilide substrates with a neutral or basic amino acid residue and dipeptide substrates such as L-leucyl-glycine. The activity of bleomycin hydrolase was inhibited by two thiol protease inhibitors, E-64 and leupeptin, as well as by N-ethylmaleimide. These results suggest that bleomycin hydrolase is a thiol aminopeptidase. Magnesium ion, sodium chloride, ethylenediaminetetraacetic acid and 1,2-dihydroxybenzene-3,5-disulfonic acid specifically activated the enzymatic hydrolysis of L-arginine- p -nitroanilide, but did not that of L-leucine- p -nitroanilide. Lineweaver-Burk plots showed that Km values of the enzymatic activity for L-arginine- p -nitroanilide were altered by these reagents, although Vmax values were almost unaltered.