title: E2 superfamily of ubiquitin-conjugating enzymes: constitutively active or activated through phosphorylation in the catalytic cleft.
abstract: Protein phosphorylation is a modification that offers a dynamic and reversible mechanism to regulate the majority of cellular processes. Numerous diseases are associated with aberrant regulation of phosphorylation-induced switches. Phosphorylation is emerging as a mechanism to modulate ubiquitination by regulating key enzymes in this pathway. The molecular mechanisms underpinning how phosphorylation regulates ubiquitinating enzymes, however, are elusive. Here, we show the high conservation of a functional site in E2 ubiquitin-conjugating enzymes. In catalytically active E2s, this site contains aspartate or a phosphorylatable serine and we refer to it as the conserved E2 serine/aspartate (CES/D) site. Molecular simulations of substrate-bound and -unbound forms of wild type, mutant and phosphorylated E2s, provide atomistic insight into the role of the CES/D residue for optimal E2 activity. Both the size and charge of the side group at the site play a central role in aligning the substrate lysine toward E2 catalytic cysteine to control ubiquitination efficiency. The CES/D site contributes to the fingerprint of the E2 superfamily. We propose that E2 enzymes can be divided into constitutively active or regulated families. E2s characterized by an aspartate at the CES/D site signify constitutively active E2s, whereas those containing a serine can be regulated by phosphorylation.
authors.affiliation: Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, 20126, Milan (Italy).
authors.forename: Ilaria
authors.initials: I
authors.lastname: Valimberti
authors.affiliation: Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, 20126, Milan (Italy).
authors.forename: Matteo
authors.initials: M
authors.lastname: Tiberti
authors.affiliation: Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, 20126, Milan (Italy).
authors.forename: Matteo
authors.initials: M
authors.lastname: Lambrughi
authors.affiliation: Cell Cycle and Cancer Unit, St. Vincent's Institute of Medical Research and The Department of Medicine, St. Vincent's Hospital, The University of Melbourne, Fitzroy, Melbourne, Victoria 3065, Australia.
authors.forename: Boris
authors.initials: B
authors.lastname: Sarcevic
authors.affiliation: Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, 20126, Milan (Italy).
authors.forename: Elena
authors.initials: E
authors.lastname: Papaleo
mesh_terms: D001665:Binding Sites; D002384:Catalysis; D004789:Enzyme Activation; D008956:Models, Chemical; D062105:Molecular Docking Simulation; D010766:Phosphorylation; D011485:Protein Binding; D025801:Ubiquitin / Q000737:chemistry* / Q000648:ultrastructure*; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry* / Q000648:ultrastructure*; D054875:Ubiquitination
chemical_list: D025801:Ubiquitin; D044763:Ubiquitin-Conjugating Enzymes