BirA is the Escherichia coli biotin ligase that site-specifically biotinylates a lysine side chain within a 15-amino acid acceptor peptide (also known as Avi-tag). As a complementary approach to in vivo biotinylation of Avi-tag-bearing proteins, we developed a protocol for producing recombinant BirA ligase for in vitro biotinylation.

Aug 9, 2024 · By fusing the biotin ligase BirA and an Avi-tag variant to the candidate E3 ligase and ubiquitin, respectively, we were able to specifically enrich bona fide substrates of a ligase using a one-step streptavidin pulldown under denaturing conditions.

Site specific biotinylation using the Avi-tag and BirA. Updated: 18.01.23. The Avi-tag is a 15aa peptide sequence (glndifeaqkiewhe) recognized by the E. coli biotin ligase BirA, which biotinylates the lysine in the sequence. The tag can be used N- or …

We describe here procedures for AviTag insertion by inverse PCR, purification of BirA fused to glutathione-S-transferase (GST-BirA) from E. coli, BirA biotinylation of purified protein, and gel-shift analysis by SDS-PAGE to quantify the extent of biotinylation.

BirA Biotin Protein Ligase for site-specific enzymatic biotinylation and tagging of proteins and antibodies. In vitro biotinylation. Expressed in E. coli. >90% Purity (SDS-PAGE, Coomassie staining) and high activity. $240.00/100 ug.

BirA has been shown to effectively biotinylate AviTag expressing proteins in bacteria, yeast, insect and mammalian cells. Check out Kay et al., 2009 for methods on biotinylating proteins with BirA. You can find plasmids that contain both AviTag and BirA at Addgene!

BirA Biotin-protein ligase (EC 6.3.4.15) activates biotin to form biotinyl 5'-adenylate and transfers the biotin to biotin-accepting proteins (such as AviTag™ peptide). It also functions as a biotin operon repressor. The protein is encoded by the birA gene.

BirA ligase specifically biotinylates certain tags on target proteins. Such tags should be added recombinantly to the protein of interest. The native E. coli BirA biotin ligase substrate is a 75-amino acid-long peptide derived from Biotin Carboxyl Carrier Protein (BCCP).

In vitro biotinylation of AviTag-fused proteins can be accomplished using purified recombinant BirA proteins, which have been tagged with GST, 6xHis and/or FLAG for convenient detection or for elimination of BirA by immunoprecipitation or …

Recombinant Escherichia coli BirA (Bifunctional ligase/repressor), full length, encompassing amino acids 2-321(end). This construct contains an N-terminal GST-tag. This recombinant protein was affinity purified.