title: Nonhydrolyzable ubiquitin-isopeptide isosteres as deubiquitinating enzyme probes.
abstract: We demonstrate that oxime ligation is an efficient, straightforward, and generally applicable strategy for generating nonhydrolyzable ubiquitin (Ub)-isopeptide isosteres. We synthesized nonhydrolyzable K48- and K63-linked Ub-isopeptide isosteres to investigate the selectivity of deubiquitinating enzymes for specific linkages employing surface plasmon resonance spectroscopy. The results indicate that deubiquitinating enzymes specifically recognize the local peptide sequence flanking Ub-branched lysine residues in target proteins. The described strategy allows the systematic investigation of sequence requirements for substrate selectivity of deubiquitinating enzymes.
authors.affiliation: Division of Cell Biology and The Netherlands Proteomics Centre, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands.
authors.forename: Anitha
authors.initials: A
authors.lastname: Shanmugham
authors.forename: Alexander
authors.initials: A
authors.lastname: Fish
authors.forename: Mark P A
authors.initials: MP
authors.lastname: Luna-Vargas
authors.forename: Alex C
authors.initials: AC
authors.lastname: Faesen
authors.forename: Farid
authors.initials: F
authors.lastname: El Oualid
authors.forename: Titia K
authors.initials: TK
authors.lastname: Sixma
authors.forename: Huib
authors.initials: H
authors.lastname: Ovaa
mesh_terms: D000595:Amino Acid Sequence; D015374:Biosensing Techniques / Q000379:methods*; D010450:Endopeptidases / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D013379:Substrate Specificity; D025801:Ubiquitin / Q000737:chemistry / Q000378:metabolism*
chemical_list: D010455:Peptides; D025801:Ubiquitin; D010450:Endopeptidases; C091939:ubiquitin isopeptidase