A HUMAN DE-UBIQUITINATING ENZYME WITH BOTH ISOPEPTIDASE AND PEPTIDASEACTIVITIES IN-VITRO

Some enzymatic and physicochemical properties of a human ubiquitin-spe cific isopeptidase are reported. The enzyme was purified to homogeneit y from red blood cells and its specificity towards polymeric ubiquitin substrates suggests a de-ubiquitinating activity capable of cleaving 'head-to-tail' polyUb chains as well as isoamide 'branched' Ub dimers. K-M values show a 10 fold preference for the cleavage of branched Ub dimers over head-to-tail Ub dimers. The enzymatic activity can be stro ngly inhibited by various peptides containing either of the cleavage s ite sequences found in Ub polymers, but not by unrelated peptides. The enzyme is monomeric under reducing conditions and exhibits a globular shape with an average diameter of 9 nm, an S-20,S-w value of 5.2 S an d a molar mass of 110 kDa +/- 10%. Because the enzyme cleaves both pep tide-linked and isopeptide-linked Ub moieties from substrates, me prop ose to name it de-ubiquitinase rather than isopeptidase.