| Autor: |
Liu J; Department of Pharmaceutical Chemistry and the Cardiovascular Research Institute , University of California San Francisco , 555 Mission Bay Boulevard South , San Francisco , California 94158 , United States., Li S; Department of Pharmaceutical Chemistry and the Cardiovascular Research Institute , University of California San Francisco , 555 Mission Bay Boulevard South , San Francisco , California 94158 , United States.; Department of Chemistry and Center for Therapeutics and Diagnostics , Georgia State University , Atlanta , Georgia 30302 , United States., Aslam NA; Hangzhou Research Institute of Technical Institute of Physics and Chemistry, Chinese Academy of Sciences , Hangzhou 310018 , China., Zheng F; Hangzhou Research Institute of Technical Institute of Physics and Chemistry, Chinese Academy of Sciences , Hangzhou 310018 , China., Yang B; Department of Pharmaceutical Chemistry and the Cardiovascular Research Institute , University of California San Francisco , 555 Mission Bay Boulevard South , San Francisco , California 94158 , United States., Cheng R; Department of Chemistry and Biochemistry , University of Delaware , Newark , Delaware 19716 , United States., Wang N; Department of Pharmaceutical Chemistry and the Cardiovascular Research Institute , University of California San Francisco , 555 Mission Bay Boulevard South , San Francisco , California 94158 , United States., Rozovsky S; Department of Chemistry and Biochemistry , University of Delaware , Newark , Delaware 19716 , United States., Wang PG; Department of Chemistry and Center for Therapeutics and Diagnostics , Georgia State University , Atlanta , Georgia 30302 , United States., Wang Q; Hangzhou Research Institute of Technical Institute of Physics and Chemistry, Chinese Academy of Sciences , Hangzhou 310018 , China., Wang L; Department of Pharmaceutical Chemistry and the Cardiovascular Research Institute , University of California San Francisco , 555 Mission Bay Boulevard South , San Francisco , California 94158 , United States. |
| Abstrakt: |
Genetically introducing covalent bonds into proteins in vivo with residue specificity is affording innovative ways for protein research and engineering, yet latent bioreactive unnatural amino acids (Uaas) genetically encoded to date react with one to few natural residues only, limiting the variety of proteins and the scope of applications amenable to this technology. Here we report the genetic encoding of (2 R)-2-amino-3-fluoro-3-(4-((2-nitrobenzyl)oxy) phenyl) propanoic acid (FnbY) in Escherichia coli and mammalian cells. Upon photoactivation, FnbY generated a reactive quinone methide (QM), which selectively reacted with nine natural amino acid residues placed in proximity in proteins directly in live cells. In addition to Cys, Lys, His, and Tyr, photoactivated FnbY also reacted with Trp, Met, Arg, Asn, and Gln, which are inaccessible with existing latent bioreactive Uaas. FnbY thus dramatically expanded the number of residues for covalent targeting in vivo. QM has longer half-life than the intermediates of conventional photo-cross-linking Uaas, and FnbY exhibited cross-linking efficiency higher than p-azido-phenylalanine. The photoactivatable and multitargeting reactivity of FnbY with selectivity toward nucleophilic residues will be valuable for addressing diverse proteins and broadening the scope of applications through exploiting covalent bonding in vivo for chemical biology, biotherapeutics, and protein engineering. |
