Anti SUMO  

Western blot of hSUMO fusion protein. Anti-SUMO antibody, generated by immunization with recombinant human SUMO, was tested by western blot against a SUMO-GFP fusion protein after cleavage by proteases. Dilution of the antibody between 1:1,000 and 1:5,000 showed strong reactivity specifically with the SUMO portion of the fusion protein ~11.5kDa (arrowhead). In this blot the antibody was used at a 1:2000 dilution incubated overnight at 4° C in 5% BLOTTO (p/n B501-0500) in TTBS. Detection occurred using a 1:2000 dilution of HRP-labeled Donkey anti-Rabbit IgG (p/n 611-703-127) for 1 hour at room temperature. A chemiluminescence system was used for signal detection (Roche). Other detection systems will yield similar results. Data contributed by M. Malakhov, www.lifesensors.com, personal communication.

Most modifiers mature by proteolytic processing from inactive precursors (a; amino acid). Arrowheads point to the cleavage sites. Ubiquitin is expressed either as polyubiquitin or as a fusion with ribosomal proteins. Conjugation requires activating (E1) and conjugating (E2) enzymes that form thiolesters (S) with the modifiers. Modification of cullins by RUB involves SCF(SKP1/cullin-1/F-box protein) /CBC(cullin-2/elongin B/elonginC) -like E3 enzymes that are also involved in ubiquitination. In contrast to ubiquitin, the UBLs do not seem to form multi-UBL chains. UCRP(ISG15) resembles two ubiquitin moieties linked head-to-tail. Whether HUB1 functions as a modifier is currently unclear. APG12 and URM1 are distinct from the other modifiers because they are unrelated in sequence to ubiquitin. Data contributed by S.Jentsch.