Human Small Ubiquitin-like Modifier 3 (SUMO3), also known as SMT3A, is synthesized as a 103 amino acid (aa), propeptide with a predicted 11.5 kDa. SUMO3 contains a two aa C-terminal prosegment. Human SUMO3 shares 83% sequence identity with mouse SUMO3. SUMO3 also has high aa sequence homology to SUMO2 and SUMO4, 87% and 75%, respectively. SUMO3 shares only 47% sequence identity with SUMO1. SUMOs are a family of small, related proteins that can be enzymatically attached to a target protein by a post-translational modification process termed SUMOylation (1-3). All SUMO proteins share a conserved Ubiquitin domain and a C-terminal diglycine cleavage/attachment site. Following prosegment cleavage, the C-terminal glycine residue of SUMO3 is enzymatically attached to a lysine residue on a target protein. In humans, SUMO3 is conjugated to a variety of molecules in the presence of the SAE1/UBA2 SUMO-activating (E1) enzyme and the UBE2I/Ubc9 SUMO-conjugating (E2) enzyme (4,5). In yeast, the SUMO-activating (E1) enzyme is Aos1/Uba2p (6). Because of the high level of sequence homology most studies report effects of SUMO2/3. For example, addition of SUMO2/3 was shown to modulate the function of ARHGAP21, a RhoGAP protein known to be involved in cell migration (7). Other reports indicate that the conjugation by SUMO2/3, but not SUMO1, may represent an important mechanism to protect neurons during episodes of cerebral ischemia (8,9). However, studies suggest that SUMO2/3 expression is regulated in an isoform-specific manner since oxidative stress downregulated the transcription of SUMO3 but not SUMO2 (10).
SUMO-3 modified with biotin via primary amine coupling results in modification of lysine residues as well as the N-terminus. Although having a fully functional C-terminus, lysine modification may limit the ability of this reagent to propagate poly-SUMO chains. Biotinylated SUMO-3 can be detected using avidin-linked reagents for higher efficiency and sensitivity than with antibodies.
