NMR‐derived solution structure of SUMO from Drosophila melanogaster (dSmt3)

Abstract

Post-translational modification of proteins by ubiquitin and ubiquitin-like proteins (UBLs) is a crucial mechanism that regulates the activities and functions of many proteins in cells. Among them, the covalent attachment of the ubiquitin molecule is the best characterized modification of substrate proteins.1,2 The protein dSmt3 belongs to this growing family of UBLs and is the Drosophila structural and functional homologue of mammalian SUMO-1 (Small Ubiquitin-like MOdifier), the most extensively characterized member among the UBLs.3,4 Most of the previous studies investigated ubiquitin and SUMO-15; however, the characterization of other SUMO family members is also mandatory for complete understanding of the functional and mechanistic aspects of SUMOylation pathway. In the past few years, the role of dSmt3 has been discovered in a number of physiologically important processes. Drosophila tramtrack 69 (Ttk69), the zinc finger transcriptional repressor of neuronal cell differentiation (playing an important role in photoreceptor cell development in the compound eye), has been identified as the substrate for dSmt3 modification.6 Ttk69 and dSmt3 proteins colocalize at polytene chromosome sites in vivo, whereas the conjugated form of Ttk69 can bind Ttk69 DNA sites in vitro. Further, the transcription factor dorsal is known to be modified with dSmt37 and is also required for Drosophila melanogaster metamorphosis.8 Lehembre et al.6 have also showed that dSmt3 is able to form conjugates with target proteins of SUMO-1 in human cells also indicating a high conservation of the Smt3 conjugation pathway. Thus, with a view to gain structural insight into dSmt3 and how it could be similar or dissimilar from mammalian SUMO and other members of SUMO family proteins in terms of structure, function, and protein–protein interaction, we undertook NMR studies of the Drosophila homologue of mammalian SUMO-1 (dSmt3) which shares 77, 75, 55, and 13.8% identity with human SUMO-3, SUMO2, SUMO-1, and ubiquitin, respectively. Here, we report the NMR-derived solution structure and surface charge topology of dSmt3 and the results are discussed in comparison with ubiquitin, human SUMO-1, -2, -3, and ySmt3 structures.