title: Adding an unnatural covalent bond to proteins through proximity-enhanced bioreactivity.
abstract: Natural proteins often rely on the disulfide bond to covalently link side chains. Here we genetically introduce a new type of covalent bond into proteins by enabling an unnatural amino acid to react with a proximal cysteine. We demonstrate the utility of this bond for enabling irreversible binding between an affibody and its protein substrate, capturing peptide-protein interactions in mammalian cells, and improving the photon output of fluorescent proteins.
authors.affiliation: The Jack H. Skirball Center for Chemical Biology and Proteomics, The Salk Institute for Biological Studies, La Jolla, California, USA.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Xiang
authors.forename: Haiyan
authors.initials: H
authors.lastname: Ren
authors.forename: Ying S
authors.initials: YS
authors.lastname: Hu
authors.forename: Irene
authors.initials: I
authors.lastname: Coin
authors.forename: Jing
authors.initials: J
authors.lastname: Wei
authors.forename: Hu
authors.initials: H
authors.lastname: Cang
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D000604:Amino Acyl-tRNA Synthetases / Q000235:genetics; D000818:Animals; D003545:Cysteine / Q000737:chemistry; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005453:Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D010649:Phenylalanine / Q000031:analogs & derivatives* / Q000737:chemistry; D017785:Photons; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D051381:Rats; D018019:Receptors, Corticotropin-Releasing Hormone / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D000099052:CRF Receptor, Type 1
chemical_list: D011506:Proteins; D018019:Receptors, Corticotropin-Releasing Hormone; D011994:Recombinant Proteins; D010649:Phenylalanine; D000099052:CRF Receptor, Type 1; D003546:Cysteine Endopeptidases; D000604:Amino Acyl-tRNA Synthetases; D003545:Cysteine
references.citation: Biochem Pharmacol. 1992 Sep 25;44(6):1201-7
references.pmid: 1417942
references.citation: MAbs. 2012 Jan-Feb;4(1):17-23
references.pmid: 22327427
references.citation: Proc Natl Acad Sci U S A. 2004 Nov 30;101(48):16745-9
references.pmid: 15556995
references.citation: Biochem J. 2005 Sep 1;390(Pt 2):455-64
references.pmid: 15901239
references.citation: Angew Chem Int Ed Engl. 2004 Sep 20;43(37):4848-70
references.pmid: 15372570
references.citation: Proc Natl Acad Sci U S A. 2001 Jul 17;98(15):8480-4
references.pmid: 11447282
references.citation: Trends Biochem Sci. 2011 Apr;36(4):229-37
references.pmid: 21055949
references.citation: Annu Rev Biochem. 2008;77:383-414
references.pmid: 18366325
references.citation: Protein Expr Purif. 2012 Mar;82(1):240-51
references.pmid: 22085722
references.citation: Nat Rev Mol Cell Biol. 2002 Nov;3(11):836-47
references.pmid: 12415301
references.citation: Proc Natl Acad Sci U S A. 2003 Mar 18;100(6):3191-6
references.pmid: 12604795
references.citation: J Biol Chem. 2009 Nov 20;284(47):32906-13
references.pmid: 19759009
references.citation: Nat Chem Biol. 2009 Aug;5(8):593-9
references.pmid: 19561621
references.citation: Mol Biosyst. 2009 Sep;5(9):931-4
references.pmid: 19668857
references.citation: Proc Natl Acad Sci U S A. 2003 Jan 7;100(1):56-61
references.pmid: 12518054
references.citation: Angew Chem Int Ed Engl. 2002 Aug 2;41(15):2840-2
references.pmid: 12203503
references.citation: Science. 2005 May 27;308(5726):1318-21
references.pmid: 15919995
references.citation: Protein Sci. 2009 Feb;18(2):460-6
references.pmid: 19165727
references.citation: Angew Chem Int Ed Engl. 2011 Aug 22;50(35):8077-81
references.pmid: 21751313
references.citation: Angew Chem Int Ed Engl. 2005 Sep 12;44(36):5788-809
references.pmid: 16149114
references.citation: Ann N Y Acad Sci. 1999 Oct 20;885:312-28
references.pmid: 10816663
references.citation: ACS Chem Biol. 2011 Jul 15;6(7):733-43
references.pmid: 21545173
references.citation: Biochem J. 2009 Mar 15;418(3):567-74
references.pmid: 19143658
references.citation: Angew Chem Int Ed Engl. 2011 Sep 5;50(37):8692-6
references.pmid: 21805546
references.citation: Nat Neurosci. 2007 Aug;10(8):1063-72
references.pmid: 17603477
references.citation: Annu Rev Biochem. 2010;79:413-44
references.pmid: 20307192