Patent ID: 8871456

Claim:
A method for preparing a protein conjugate, the method comprising: contacting a fusion protein with a lipoic acid analog in the presence of a lipoic acid ligase polypeptide to produce a protein conjugate in which the lipoic acid analog is linked to the fusion protein, wherein: the lipoic acid analog is a substrate of the lipoic acid ligase polypeptide and comprises either a functional group handle or a directly detectable moiety, the fusion protein comprises the target protein and an acceptor polypeptide, the acceptor polypeptide consists of the amino acid sequence Xaa 1 Xaa 2 Xaa 3 Xaa 4 Xaa 5 Xaa 6 Xaa 7 Xaa 8 Xaa 9 Lys Xaa 10 Xaa 11 Xaa 12 Xaa 13 Xaa 14 or a fragment thereof having at least 12 amino acid residues, each of Xaa 3 , Xaa 6 and Xaa 14 , independently, being a hydrophobic amino acid residue, each of Xaa 1 , Xaa 2 , Xaa 4 , Xaa 5 , Xaa 8 , Xaa 10 , Xaa 11 , and Xaa 12 , independently, being any amino acid residue, and each of Xaa 7 , Xaa 9 , and Xaa 13 , independently, being either E or D; and the lipoic acid ligase polypeptide comprises the amino acid sequence of a wild-type lipoic acid ligase, or is a functional mutant of a wild-type lipoic acid ligase, which has an amino acid sequence at least 85% identical to SEQ ID NO:11, has lipoic acid ligase activity, and comprises an amino acid residue substitution at the position corresponding to W37 in SEQ ID NO:11.