Chemical techniques for protein biotinylation lack reaction specificity and frequently inactivate the function of the target proteins. Creative Biolabs has developed a unique protein and antibody site-specific biotinylation platform named as BtAP-Tag® that well eliminates these issues.
In Creative Biolabs, we have many years of experience in standard antibody conjugation from organic dyes and protein conjugation to other dyes, enzymes or other biomolecules. Labeling of other functional groups, special purification of conjugates and bioassays may also be available.
On this platform, a 15-amino acid sequence (the Biotin acceptor peptide, BtAP) is genetically fused with the target protein of interest. The BtAP serves as an in vivo substrate for E. coli biotin holoenzyme synthetase (BirA), an enzyme that naturally select biotinylates E. coli biotin carboxyl carrier protein (BCCP). We have developed an efficient bacterial expression vector that allows expression of the target protein fused with BtAP and expression of the BirA gene under a bacterial operon. Using this vector and an optimized bacterial host strain, the BtAP-fused target protein and BirA can be co-expressed and the target protein will be fully biotinylated.
No matter the BtAP is fused on the N-terminus, C-terminus or inserted into loops of the target protein, it can be efficiently biotinylated. Biotinylation by this platform allows sensitive recognition by avidin or streptavidin that is conjugated to a range of biophysical probes.
We have also established a mammalian in vivo protein biotinylation platform to produce BtAP fused with biotinylated proteins in mammalian cells.