Biotin ligase (BirA), also known as biotin acetyl-CoA synthetase, is a protease widely present in prokaryotes and eukaryotes. The biotin ligase encoded by the E. coli BirA gene recognizes a specific sequence of a protein molecule and biotinylates the protein. Biotin ligase (BirA) is an indispensable enzyme for organic life to acquire biotin. The BirA receptor-polypeptide system has ultra-high specificity when biotinylated polypeptide substrates. BirA recognizes a polypeptide sequence (biotin receptor tag BAT). When the BAT-fused protein is co-expressed with BirA, the BAT sequence can be efficiently biotinylated. The biotinylated protein can be further purified by avidin or streptavidin with high affinity.