A human de-ubiquitinating enzyme with both isopeptidase and peptidase activities in vitro.
Some enzymatic and physicochemical properties of a human ubiquitin-specific isopeptidase are reported. The enzyme was purified to homogeneity 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. KM values show a 10 fold preference for the cleavage of branched Ub dimers over head-to-tail Ub dimers. The enzymatic activity can be strongly inhibited by various peptides containing either of the cleavage site 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 S20,w value of 5.2 S and a molar mass of 110 kDa +/- 10%. Because the enzyme cleaves both peptide-linked and isopeptide-linked Ub moieties from substrates, we propose to name it de-ubiquitinase rather than isopeptidase.
Department of Medical Biochemistry, Faculty of Medicine, University of Geneva, Switzerland.
L
L
Falquet
N
N
Paquet
S
S
Frutiger
G J
GJ
Hughes
K
K
Hoang-Van
J C
JC
Jaton
D000595:Amino Acid Sequence; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D010450:Endopeptidases / Q000096:biosynthesis* / Q000737:chemistry; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D013379:Substrate Specificity; D014452:Ubiquitins / Q000737:chemistry / Q000378:metabolism*
D046911:Macromolecular Substances; D014452:Ubiquitins; D010450:Endopeptidases; C091939:ubiquitin isopeptidase