title: Linkage via K27 Bestows Ubiquitin Chains with Unique Properties among Polyubiquitins.
abstract: Polyubiquitination, a critical protein post-translational modification, signals for a diverse set of cellular events via the different isopeptide linkages formed between the C terminus of one ubiquitin (Ub) and the ɛ-amine of K6, K11, K27, K29, K33, K48, or K63 of a second Ub. We assembled di-ubiquitins (Ub2) comprising every lysine linkage and examined them biochemically and structurally. Of these, K27-Ub2 is unique as it is not cleaved by most deubiquitinases. As this remains the only structurally uncharacterized lysine linkage, we comprehensively examined the structures and dynamics of K27-Ub2 using nuclear magnetic resonance, small-angle neutron scattering, and in silico ensemble modeling. Our structural data provide insights into the functional properties of K27-Ub2, in particular that K27-Ub2 may be specifically recognized by K48-selective receptor UBA2 domain from proteasomal shuttle protein hHR23a. Binding studies and mutagenesis confirmed this prediction, further highlighting structural/recognition versatility of polyubiquitins and the potential power of determining function from elucidation of conformational ensembles.
authors.affiliation: Department of Chemistry and Biochemistry, Center for Biomolecular Structure and Organization, University of Maryland, College Park, MD 20742, USA; Departments of Biology & Chemistry, Syracuse University, Syracuse, NY 13244, USA. Electronic address: cacastan@syr.edu.
authors.forename: Carlos A
authors.initials: CA
authors.lastname: Castañeda
authors.affiliation: Department of Chemistry and Biochemistry, Center for Biomolecular Structure and Organization, University of Maryland, College Park, MD 20742, USA.
authors.forename: Emma K
authors.initials: EK
authors.lastname: Dixon
authors.affiliation: Institut des Sciences Analytiques, UMR5280-Université de Lyon, 69100 Villeurbanne, France.
authors.forename: Olivier
authors.initials: O
authors.lastname: Walker
authors.affiliation: Department of Chemistry and Biochemistry, Center for Biomolecular Structure and Organization, University of Maryland, College Park, MD 20742, USA.
authors.forename: Apurva
authors.initials: A
authors.lastname: Chaturvedi
authors.affiliation: Department of Chemistry and Biochemistry, Center for Biomolecular Structure and Organization, University of Maryland, College Park, MD 20742, USA.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Nakasone
authors.affiliation: NIST Center for Neutron Research, National Institute of Standards and Technology, Gaithersburg, MD 20899-8562, USA.
authors.forename: Joseph E
authors.initials: JE
authors.lastname: Curtis
authors.affiliation: Department of Chemistry, Virginia Commonwealth University, Richmond, VA 23284, USA.
authors.forename: Megan R
authors.initials: MR
authors.lastname: Reed
authors.affiliation: NIST Center for Neutron Research, National Institute of Standards and Technology, Gaithersburg, MD 20899-8562, USA.
authors.forename: Susan
authors.initials: S
authors.lastname: Krueger
authors.affiliation: Department of Chemistry, Virginia Commonwealth University, Richmond, VA 23284, USA.
authors.forename: T Ashton
authors.initials: TA
authors.lastname: Cropp
authors.affiliation: Department of Chemistry and Biochemistry, Center for Biomolecular Structure and Organization, University of Maryland, College Park, MD 20742, USA. Electronic address: fushman@umd.edu.
authors.forename: David
authors.initials: D
authors.lastname: Fushman
mesh_terms: D001665:Binding Sites; D045643:DNA Repair Enzymes / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008239:Lysine / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D025821:Polyubiquitin / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D053838:Scattering, Small Angle; D054875:Ubiquitination
chemical_list: D004268:DNA-Binding Proteins; D025821:Polyubiquitin; C086985:RAD23A protein, human; D045643:DNA Repair Enzymes; D008239:Lysine