Ubiquitin Activation (E1, E2 Enzymes) Antibody Sampler Kit

Background

Ubiquitin is a conserved polypeptide unit that plays an important role in the ubiquitin-proteasome pathway. Ubiquitin can be covalently linked to many cellular proteins for degradation by the 26S proteasome. Three components are involved in the target protein-ubiquitin conjugation process. Ubiquitin is first activated by forming a thioester complex with the ubiquitin-activating enzyme (UBE1 or E1). The activated ubiquitin is subsequently transferred to the ubiqutin-carrier protein (conjugating enzyme) E2, and then from E2 to ubiqutin ligase E3 for final delivery to the epsilon-NH₂ of the target protein lysine residue (1-3). The ubiquitin-proteasome pathway has been implicated in a wide range of normal biological processes and in disease-related abnormalities. Several proteins such as IκB, p53, cdc25a, and Bcl-2 have been shown to be targets for the ubiquitin-proteasome process as part of the regulation of cell cycle progression, differentiation, cell stress response, and apoptosis (4-7). UBC3, the mammalian ortholog of yeast cdc34, and UBC3B, a UBC3 family member, are E2 ubiquitin-carrier proteins. UBC3, in concert with SCF-Skp2 (Skp1, Cullin, F-box protein/Skp2) complex, mediates cell cycle progression from G1 to S phase by targeting the CDK inhibitor p27 for proteolysis (8). UBC3B, in concert with SCFb-Trcp (Skp1, Cullin and F-box protein/b-Trcp) complex, mediates degradation of β-catenin (9). UbcH5C is a universally expressed E2 ubiquitin conjugating enzyme and member of the UbcH5 family that also includes UbcH5A and UbcH5B (10). Evidence suggests that UbcH5 plays an important role in regulating a number of signaling pathways by catalyzing the ubiquitination of key target proteins, including p53, PCNA, the IκB kinase proein NEMO, and the apoptosis inhibitor BRUCE (11-14). UBE2L3, also commonly referred to as UBCH7, is a ubiquitin-conjugating enzyme that has been linked to the ubiquitination of numerous substrates via its interaction with protein-ubiquitin E3 ligases, such as NEDD4 (15), E6AP (16), Parkin (17), c-Cbl (18), and Triad1 (19,20). UBE2N/Ubc13 is a ubiquitin-E2-conjugating enzyme that catalyzes K63-linked polyubiquitin chain formation (21,22). UBE2N forms a heterodimer with MMS2 or Uev1A to exert its E2 ligase function. The UBE2N/MMS2 and UBE2N/Uev1A heterodimers catalyze different modes of target protein ubiquitination to mediate various signaling pathways (23-25) including DNA damage and recombination, p53 and check point control, cell cycle (26-30), immunoreceptor signaling (31,32), and endocytosis (33).