An immobilized biotin ligase: surface display of Escherichia coli BirA on Saccharomyces cerevisiae.
The Escherichia coli biotin ligase enzyme BirA has been extensively used in recent years to generate site-specifically biotinylated proteins via a biotin acceptor peptide tag. In the present study, BirA was displayed for the first time on the yeast Saccharomyces cerevisiae using the Aga1p-Aga2p platform and assayed using a peptide-tagged protein as the substrate. The enzyme is fully functional and resembles the soluble form in many of its properties, but the yeast-displayed enzyme demonstrates stability and reusability on the time scale of weeks. Thus, the yeast-displayed BirA system represents a facile and highly economical alternative for producing site-specifically biotinylated proteins.
Parthasarathy
Ranganath
R
Department of Chemical and Biomolecular Engineering and Laboratory for Research on the Structure of Matter, University of Pennsylvania, 220 South 33rd Street, Philadelphia, 19104, USA.
Bajaj
Jitin
J
Boder
Eric T
ET
D000595:Amino Acid Sequence; D001709:Biotechnology; D001710:Biotin / Q000378:metabolism; D019731:Carbon-Nitrogen Ligases / Q000235:genetics* / Q000378:metabolism*; D002384:Catalysis; D002462:Cell Membrane / Q000201:enzymology; D003001:Cloning, Molecular; D004800:Enzymes, Immobilized; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism*
D004800:Enzymes, Immobilized; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D001710:Biotin; D019731:Carbon-Nitrogen Ligases; C058125:birA protein, E coli