title: Two rapid catalyst-free click reactions for in vivo protein labeling of genetically encoded strained alkene/alkyne functionalities.
abstract: Detailed kinetic analyses of inverse electron-demand Diels–Alder cycloaddition and nitrilimine-alkene/alkyne 1,3-diploar cycloaddition reactions were conducted and the reactions were applied for rapid protein bioconjugation. When reacted with a tetrazine or a diaryl nitrilimine, strained alkene/alkyne entities including norbornene, trans-cyclooctene, and cyclooctyne displayed rapid kinetics. To apply these “click” reactions for site-specific protein labeling, five tyrosine derivatives that contain a norbornene, trans-cyclooctene, or cyclooctyne entity were genetically encoded into proteins in Escherichia coli using an engineered pyrrolysyl-tRNA synthetase-tRNA(CUA)(Pyl) pair. Proteins bearing these noncanonical amino acids were successively labeled with a fluorescein tetrazine dye and a diaryl nitrilimine both in vitro and in living cells.
authors.affiliation: Department of Chemistry, Texas A&M University , College Station, Texas 77843, United States.
authors.forename: Yadagiri
authors.initials: Y
authors.lastname: Kurra
authors.forename: Keturah A
authors.initials: KA
authors.lastname: Odoi
authors.forename: Yan-Jiun
authors.initials: YJ
authors.lastname: Lee
authors.forename: Yanyan
authors.initials: Y
authors.lastname: Yang
authors.forename: Tongxiang
authors.initials: T
authors.lastname: Lu
authors.forename: Steven E
authors.initials: SE
authors.lastname: Wheeler
authors.forename: Jessica
authors.initials: J
authors.lastname: Torres-Kolbus
authors.forename: Alexander
authors.initials: A
authors.lastname: Deiters
authors.forename: Wenshe R
authors.initials: WR
authors.lastname: Liu
mesh_terms: D000475:Alkenes / Q000737:chemistry*; D000480:Alkynes / Q000737:chemistry*; D002384:Catalysis; D057930:Click Chemistry; D034242:Cyclooctanes / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000737:chemistry; D007700:Kinetics; D008958:Models, Molecular; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry*; D013194:Staining and Labeling / Q000379:methods*; D014443:Tyrosine / Q000737:chemistry
chemical_list: D000475:Alkenes; D000480:Alkynes; D034242:Cyclooctanes; D029968:Escherichia coli Proteins; D011506:Proteins; D014443:Tyrosine