Insights into E3 ligase activity revealed by a SUMO-RanGAP1-Ubc9-Nup358 complex

Structural Biology Program, Sloan-Kettering Institute, New York, New York 10021, USA.
Nature (Impact Factor: 41.46). 07/2005; 435(7042):687-92. DOI: 10.1038/nature03588
Source: PubMed


SUMO-1 (for small ubiquitin-related modifier) belongs to the ubiquitin (Ub) and ubiquitin-like (Ubl) protein family. SUMO conjugation occurs on specific lysine residues within protein targets, regulating pathways involved in differentiation, apoptosis, the cell cycle and responses to stress by altering protein function through changes in activity or cellular localization or by protecting substrates from ubiquitination. Ub/Ubl conjugation occurs in sequential steps and requires the concerted action of E2 conjugating proteins and E3 ligases. In addition to being a SUMO E3, the nucleoporin Nup358/RanBP2 localizes SUMO-conjugated RanGAP1 to the cytoplasmic face of the nuclear pore complex by means of interactions in a complex that also includes Ubc9, the SUMO E2 conjugating protein. Here we describe the 3.0-A crystal structure of a four-protein complex of Ubc9, a Nup358/RanBP2 E3 ligase domain (IR1-M) and SUMO-1 conjugated to the carboxy-terminal domain of RanGAP1. Structural insights, combined with biochemical and kinetic data obtained with additional substrates, support a model in which Nup358/RanBP2 acts as an E3 by binding both SUMO and Ubc9 to position the SUMO-E2-thioester in an optimal orientation to enhance conjugation.

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Available from: David Reverter, Jan 22, 2014
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    • "The archetypal consensus SIM consists of four hydrophobic amino acids adjacent to a cluster of negatively charged residues [3]-[8]. Structural studies revealed that this SIM forms an extended β-strand that inserts between the α-helix and second β-strand in all three SUMO paralogs [4], [7]–[10]. However, little is understood regarding how proteins preferentially recognize SUMO-1 or SUMO-2, although for some SIM containing proteins the cluster of negatively charged residues confers binding specificity for SUMO-1 over SUMO-2 [4], [11]. "
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    ABSTRACT: SUMO-binding proteins interact with SUMO modified proteins to mediate a wide range of functional consequences. Here, we report the identification of a new SUMO-binding protein, ZNF261. Four human proteins including ZNF261, ZNF198, ZNF262, and ZNF258 contain a stretch of tandem zinc fingers called myeloproliferative and mental retardation (MYM)-type zinc fingers. We demonstrated that MYM-type zinc fingers from ZNF261 and ZNF198 are necessary and sufficient for SUMO-binding and that individual MYM-type zinc fingers function as SUMO-interacting motifs (SIMs). Our binding studies revealed that the MYM-type zinc fingers from ZNF261 and ZNF198 interact with the same surface on SUMO-2 recognized by the archetypal consensus SIM. We also present evidence that MYM-type zinc fingers in ZNF261 contain zinc, but that zinc is not required for SUMO-binding. Immunofluorescence microscopy studies using truncated fragments of ZNF198 revealed that MYM-type zinc fingers of ZNF198 are necessary for localization to PML-nuclear bodies (PML-NBs). In summary, our studies have identified and characterized the SUMO-binding activity of the MYM-type zinc fingers in ZNF261 and ZNF198.
    PLoS ONE 08/2014; 9(8):e105271. DOI:10.1371/journal.pone.0105271 · 3.23 Impact Factor
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    • "Indeed, a recent structural study revealed how the second repeat increased SUMO1 binding, as well as SUMO1-specific E3 ligase activity for the RanBP2 complex (Gareau et al. 2012). As previously observed with the minimal catalytic domain of SP-RING SUMO ligases, no substrate-binding domain could be identified in the minimal catalytic fragment of RanBP2 (Pichler et al. 2002, 2004; Reverter and Lima 2005). However, RanBP2's catalytic entity in context of the full-length protein is flanked by binding sites for transport receptors, suggesting transport cargos as putative substrates. "
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    ABSTRACT: Posttranslational modification of proteins by the small ubiquitin-like modifier (SUMO) is a potent regulator of various cellular events. Hundreds of substrates have been identified, many of them involved in vital processes like transcriptional regulation, signal transduction, protein degradation, cell cycle regulation, DNA repair, chromatin organization, and nuclear transport. In recent years, protein sumoylation increasingly attracted attention, as it could be linked to heart failure, cancer, and neurodegeneration. However, underlying mechanisms involving how modification by SUMO contributes to disease development are still scarce thus necessitating further research. This review aims to critically discuss currently available concepts of the SUMO pathway, thereby highlighting regulation in the healthy versus diseased organism, focusing on neurologic aspects. Better understanding of differential regulation in health and disease may finally allow to uncover pathogenic mechanisms and contribute to the development of disease-specific therapies.
    Neuromolecular medicine 08/2013; 15(4). DOI:10.1007/s12017-013-8258-6 · 3.68 Impact Factor
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    • "NUP358 plays an important role in nucleocytoplasmic transport, being the site of interaction with Ran (the small GTPase that regulates nucleocytoplasmic transport by proteins of the karyopherin β family [47]), SUMO-modified RanGAP (the mammalian RanGTPase-activating protein that is highly concentrated at the cytoplasmic side of the nuclear pore complex [48,49]), and the export receptor CRM1 [50]. Nup358 has also been suggested to act as a platform to recruit import receptors to pre-bound cargos, as cellular substrates DBC-1 and DMAP-1 have been shown to directly bind to NUP358 and be transported by importin α/β [51]. "
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    ABSTRACT: Lentiviruses such as HIV-1 can be distinguished from other retroviruses by the cyclophilin A-binding loop in their capsid and their ability to infect non-dividing cells. Infection of non-dividing cells requires transport through the nuclear pore but how this is mediated is unknown. Here we present the crystal structure of the N-terminal capsid domain of HIV-1 in complex with the cyclophilin domain of nuclear pore protein NUP358. The structure reveals that HIV-1 is positioned to allow single-bond resonance stabilisation of exposed capsid residue P90. NMR exchange experiments demonstrate that NUP358 is an active isomerase, which efficiently catalyzes cis-trans isomerization of the HIV-1 capsid. In contrast, the distantly related feline lentivirus FIV can bind NUP358 but is neither isomerized by it nor requires it for infection. Isomerization by NUP358 may be preserved by HIV-1 to target the nuclear pore and synchronize nuclear entry with capsid uncoating.
    Retrovirology 07/2013; 10(1):81. DOI:10.1186/1742-4690-10-81 · 4.19 Impact Factor
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