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  • br E intracellular localization br


    E1 intracellular localization
    Regulation of E1 by other post-translational modifications
    Concluding remarks
    Acknowledgments We apologize to those whose work was not included because of space considerations or whose papers were unintentionally omitted. We thank Dr Peter Bullock (Tufts University) and members of the Archambault and Melendy laboratories for critical reading of this manuscript. Work in the authors’ laboratories is supported by grants from the US National Institutes of Health (NIH - AI095632), Canadian Institutes for Health Research (CIHR), Canadian Cancer Society Research Institute (CCSRI) and the Cancer Research Society (CRS).
    Introduction Sumoylation is an essential transient posttranslational modification that is predominantly detected in the nucleus and has key functions in many cellular pathways, including transcription, chromatin regulation, DNA replication, DNA damage responses, RNA splicing, Eptifibatide regulation, protein degradation, and intracellular trafficking (Droescher, Chaugule, & Pichler, 2013; Flotho & Melchior, 2013; Zhao, 2018). In humans, several different small ubiquitin-related modifier (SUMO) paralogs can be conjugated to cellular proteins. The human genome codes for five SUMO paralogs (SUMO1–5); of these, SUMO1 and the almost identical SUMO2 and SUMO3 are ubiquitously expressed. Sumoylation is executed by an enzymatic triad that covalently attaches SUMO to selected substrates in a hierarchical process. First, the unique heterodimeric SUMO E1 enzyme Aos1/Uba2 activates SUMO by ATP-driven adenylation of the SUMO C-terminus followed by formation of an energy-rich thioester bond between a Uba2 cysteine and the SUMO C-terminus. Next, SUMO is transferred to the unique E2 enzyme Ubc9 again resulting in a thioester linkage (~). Eventually, the SUMO C-terminus is conjugated to a substrate lysine residue, forming an isopeptide bond (*). The final conjugation step usually involves E3 ligases, which stabilize the interaction of the SUMO~charged E2 enzyme with the substrate. However, exceptional cases allow efficient modification also in the absence of E3 ligases (Pichler, Fatouros, Lee, & Eisenhardt, 2017). Generally, substrates can be modified at single or multiple lysines either with a single SUMO moiety or with poly SUMO chains. The most abundant sumoylation sites are acceptor lysines embedded in a SUMO consensus motif (SCM) ψKxE (ψ: hydrophobic amino acid with preference for V and I), but non-SCM lysines can also be modified especially upon stress conditions (Hendriks et al., 2017, Hendriks et al., 2018; Hendriks & Vertegaal, 2016). The number of known SUMO-ligating enzymes is very limited, with far fewer than the number of corresponding enzymes used for ubiquitin-protein ligation. In mammals, these SUMO pathway enzymes comprise the single E1 and E2 enzymes and a handful E3 ligases that belong to three different classes: 1. the SP-RING family consisting of PIAS1, PIAS2, PIAS3, PIAS4, and MMS21; 2. RanBP2; and 3. the ZNF451-family presented by ZNF451–1, ZNF451–2, ZNF451–3, and the primate-specific KIAA1586 protein (Cappadocia, Pichler, & Lima, 2015; Eisenhardt et al., 2015; Kahyo, Nishida, & Yasuda, 2001; Pichler, Gast, Seeler, Dejean, & Melchior, 2002; Sachdev et al., 2001). This small enzyme number is especially surprising in the light of thousands of SUMO substrates which have been identified in cells (Hendriks et al., 2017, Hendriks et al., 2018; Hendriks & Vertegaal, 2016). Detailed biochemical and structural analyses of the three known classes of bona fide SUMO E3 ligases revealed that specific donor-SUMO (SUMOD) positioning is a hallmark of E3-dependent catalysis along with the ability to bind E2 (Cappadocia et al., 2015; Eisenhardt et al., 2015; Reverter & Lima, 2005; Streich & Lima, 2016; Yunus & Lima, 2009). SUMOD is the SUMO that forms the thioester linkage with the E2 enzyme and its positioning leads to an optimal orientation, the so-called closed conformation, for nucleophilic attack of the incoming substrate lysine ɛ-amino group, resulting in efficient isopeptide formation. Thus, SUMOD positioning and rapid discharge of the SUMOD~E2 are diagnostic features of E3 ligases that can be monitored in in vitro sumoylation reactions.