Site-specific incorporation of redox active amino acids into proteins

A composition comprising a protein, wherein the protein comprises one or more redox active amino acids selected from the group consisting of:

• a 3,4-dihydroxy-L-phenyalanine (DHP), a 3,4,5-trihydroxy-L-phenylalanine, a 3-nitro-tyrosine, and a 3-thiol-tyrosine;
• the composition further comprising:
• i) at least one orthogonal tRNA (O-tRNA),
• ii) at least one orthogonal aminoacyl-tRNA synthetase (O-RS) comprising a Leu amino acid in a position of the O-RS corresponding to Tyr32 of SEQ ID NO:
• 4, a Ser amino acid residue in a position of the O-RS corresponding to Ala67 of SEQ ID NO:
• 4, an Asn amino acid residue in a position of the O-RS corresponding to His 70 of SEQ ID NO:
• 4, and a Gln residue in a position of the O-RS corresponding to Ala167 of SEQ ID NO:
• 4, and SEQ ID NO:
• 4 is the wild type sequence;
• wherein the O-RS comprises at least 90% identity to the RS of SEQ ID NO:
• 1 wherein the O-RS
• preferentially aminoacylates the O-tRNA with one or more of the redox active amino acids;
• and
• iii) a nucleic acid that encodes the protein, wherein the nucleic acid comprises at least one selector codon that is recognized by the O-tRNA;
• wherein the protein is a product of a translation involving the nucleic acid, the one or more redox amino acids, the O-tRNA and the O-RS.