Ubiquitin modification of substrate proteins is achieved by the activity of E1 activating, E2 conjugating and E3 ligase enzymes. Substrate proteins can be modified with one (mono-ubiquitination) or many (poly-ubiquitination) molecules of ubiquitin, and each type of modification has distinct regulatory fates.
In addition, many E3s have been implicated in human disease and are attractive targets for drug discovery. However, currently no small molecule modulators for this class of enzymes have reached the clinic. Each E3 enzyme targets a small number of proteins for Ub modification but the exact substrates are mostly unknown and their identification continues to be a challenge. E3 ligase enzymes are a large (> 500) and complex super-family, many of which contain binding domains to interact with ubiquitin, E2 enzymes and substrate proteins. In addition to substrate ubiquitination, many E3 ligases can also self- or auto-ubiquitinate in the presence of an E2 enzyme, a property that may be used as an auto-regulatory mechanism to control its own intracellular levels. In general, the detailed molecular mechanism, stoichiometries and linkage site selection of E3 enzymes are poorly understood. As with ubiquitin E3 ligases, similar activities are also part of the final conjugation processes for other UBL proteins. There are E3 enzymes that are specific for targets that are modified SUMO, NEDD8, ISG15 and presumably also for FAT10 and UFM1.