Adding an unnatural covalent bond to proteins through proximity-enhanced bioreactivity

• Published in: Nature methods Vol. 10; no. 9; pp. 885 - 888
• Main Authors: Xiang, Zheng, Ren, Haiyan, Hu, Ying S, Coin, Irene, Wei, Jing, Cang, Hu, Wang, Lei
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01.09.2013; Nature Publishing Group
• Subjects: 631/1647/338/469; 631/1647/666; 631/45/612; 631/92/96; Amino Acid Sequence; Amino acids; Amino Acyl-tRNA Synthetases - genetics; Animals; Bioinformatics; Biological Microscopy; Biological Techniques; Biomedical Engineering/Biotechnology; Biomedical research; brief-communication; Cells; Cellular proteins; Chemical bonds; Cysteine - chemistry; Cysteine Endopeptidases - chemistry; Cysteine Endopeptidases - genetics; Cysteine Endopeptidases - metabolism; Escherichia coli - genetics; Fluorescence; Life Sciences; Mammals; Molecular Sequence Data; Mutation; Phenylalanine - analogs & derivatives; Phenylalanine - chemistry; Photons; Physiological aspects; Protein binding; Protein Conformation; Protein Engineering - methods; Proteins; Proteins - chemistry; Proteins - genetics; Proteins - immunology; Proteins - metabolism; Proteomics; Rats; Receptors, Corticotropin-Releasing Hormone - genetics; Receptors, Corticotropin-Releasing Hormone - metabolism; Recombinant Proteins - genetics; Recombinant Proteins - immunology; United States
• ISSN: 1548-7091; 1548-7105
• DOI: 10.1038/nmeth.2595

An unnatural amino acid that forms a covalent bond with a proximal cysteine can be introduced into proteins, facilitating a variety of applications. 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.