title: A Central Cysteine Residue Is Essential for the Thermal Stability and Function of SUMO-1 Protein and SUMO-1 Peptide-Protein Conjugates.
abstract: SUMOylation constitutes a major post-translational modification (PTM) used by the eukaryote cellular machinery to modulate protein interactions of the targeted proteins. The small ubiquitin-like modifier-1 (SUMO-1) features a central and conserved cysteine residue (Cys52) that is located in the hydrophobic core of the protein and in tight contact with Phe65, suggesting the occurrence of an S/π interaction. To investigate the importance of Cys52 on SUMO-1 thermal stability and biochemical properties, we produced by total chemical synthesis SUMO-1 or SUMO-1 Cys52Ala peptide-protein conjugates featuring a native isopeptidic bond between SUMO-1 and a peptide derived from p53 tumor suppressor protein. The Cys52Ala modification perturbed SUMO-1 secondary structure and resulted in a dramatic loss of protein thermal stability. Moreover, the cleavage of the isopeptidic bond by the deconjugating enzyme Upl1 was significantly less efficient than for the wild-type conjugate. Similarly, the in vitro SUMOylation of RanGap1 by E1/E2 conjugating enzymes was significantly less efficient with the SUMO-1 C52A analog compared to wild-type SUMO-1. These data demonstrate the critical role of Cys52 in maintaining SUMO-1 conformation and function and the importance of keeping this cysteine intact for the study of SUMO-1 protein conjugates.
authors.affiliation: M3T-Mechanisms of Tumorigenesis and Target Therapies, Université de Lille, CNRS, Institut Pasteur de Lille, UMR 8161, F-59000 Lille, France.
authors.forename: Hervé
authors.initials: H
authors.lastname: Drobecq
authors.affiliation: M3T-Mechanisms of Tumorigenesis and Target Therapies, Université de Lille, CNRS, Institut Pasteur de Lille, UMR 8161, F-59000 Lille, France.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Boll
authors.affiliation: M3T-Mechanisms of Tumorigenesis and Target Therapies, Université de Lille, CNRS, Institut Pasteur de Lille, UMR 8161, F-59000 Lille, France.
authors.forename: Magalie
authors.initials: M
authors.lastname: Sénéchal
authors.affiliation: M3T-Mechanisms of Tumorigenesis and Target Therapies, Université de Lille, CNRS, Institut Pasteur de Lille, UMR 8161, F-59000 Lille, France.
authors.forename: Rémi
authors.initials: R
authors.lastname: Desmet
authors.affiliation: Université de Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019-UMR 8204, F-59000 Lille, France.
authors.forename: Jean-Michel
authors.initials: JM
authors.lastname: Saliou
authors.affiliation: Sorbonne Universités, UPMC Université Paris 06, École Normale Supérieure, PSL Research University, CNRS UMR 7203 LBM, F-75005, Paris, France.
authors.forename: Jean-Jacques
authors.initials: JJ
authors.lastname: Lacapère
authors.affiliation: M3T-Mechanisms of Tumorigenesis and Target Therapies, Université de Lille, CNRS, Institut Pasteur de Lille, UMR 8161, F-59000 Lille, France.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Mougel
authors.affiliation: M3T-Mechanisms of Tumorigenesis and Target Therapies, Université de Lille, CNRS, Institut Pasteur de Lille, UMR 8161, F-59000 Lille, France.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Vicogne
authors.affiliation: M3T-Mechanisms of Tumorigenesis and Target Therapies, Université de Lille, CNRS, Institut Pasteur de Lille, UMR 8161, F-59000 Lille, France.
authors.forename: Oleg
authors.initials: O
authors.lastname: Melnyk
mesh_terms: D000595:Amino Acid Sequence; D017124:Conserved Sequence; D003545:Cysteine; D006801:Humans; D008958:Models, Molecular; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism*; D000072417:Protein Domains; D055550:Protein Stability; D025842:SUMO-1 Protein / Q000737:chemistry* / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013696:Temperature; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry / Q000378:metabolism
chemical_list: D010446:Peptide Fragments; D025842:SUMO-1 Protein; D016159:Tumor Suppressor Protein p53; D003545:Cysteine