**2. Enzymes in the Ubiquitin System**

Ubiquitination is carried out by a cascade of enzymatic reactions. E1 activating enzyme uses ATP to activate the C-terminal carboxyl group of ubiquitin, which then forms a thioester bond with the cysteine residue in the active site of an E2 ubiquitin conjugating enzyme. In the final step, an E3 ubiquitin ligase transfers the ubiquitin from the E2-Ub to a specific substrate. Resulted from the serial reactions, an isopeptide bond is formed between the carboxyl group of ubiquitin and the ε-amino group of a lysine residue of the substrate protein [3].

The human genome has two E1 activating enzymes, about 40 E2 conjugating enzymes and over 600 E3 ubiquitin ligases [7]. E1 serves to distinguish ubiquitin from other ubiquitin-like proteins [8]. E2s are small proteins of about 150 amino acids, whose selectivity is generally realized via their interactions with di fferent E3s. The diverse array of E3s are categorized into three classes based on their (i) catalytic domain and (ii) the mechanism of ubiquitination reaction. The most common type of E3s is the really interesting new gene (RING) finger containing E3 ubiquitin ligases that can simultaneously bind to both the substrate and E2-Ub and directly transfer ubiquitin from the E2 to the substrate. The HECT (homologous to the E6AP carboxyl terminus) domain containing E3s and RING-between-RING (RBR) type E3s both transfer ubiquitin from E2 to form E3-Ub and then to the substrate in a two-step fashion [7,9].

Deubiquitinases (DUBs) are enzymes that remove ubiquitin from the substrate proteins. There are about 100 DUBs in humans that are classified into seven families based on their evolutionary conservation. Six of the DUB families are cysteine proteases and the last one is Zn-dependent metalloproteases [10,11].
