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In Vivo Biotinylation of Antigens in E. coli

  • Susanne Gräslund
  • Pavel Savitsky
  • Susanne Müller-Knapp
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1586)

Abstract

Site-specific biotinylation of proteins is often the method of choice to enable efficient immobilization of a protein on a surface without interfering with protein folding. The tight interaction of biotin and streptavidin is frequently used to immobilize an antigen during phage display selections of binders. Here we describe a method of in vivo biotinylation of proteins during expression in E. coli, by tagging the protein with the short biotin acceptor peptide sequence, Avi tag, and co-expression of the E. coli biotin ligase (BirA) resulting in precise biotinylation of a specific lysine residue in the tag.

Key words

Biotinylation BirA Avi-tag Antigen capturing Antigen immobilization Streptavidin IMAC SEC 

Notes

Acknowledgments

The SGC is a registered charity (number 1097737) that receives funds from AbbVie, Bayer Pharma AG, Boehringer Ingelheim, Canada Foundation for Innovation, Eshelman Institute for Innovation, Genome Canada, Innovative Medicines Initiative (EU/EFPIA) [ULTRA-DD grant no. 115766], Janssen, Merck & Co., Novartis Pharma AG, Ontario Ministry of Economic Development and Innovation, Pfizer, São Paulo Research Foundation-FAPESP, Takeda, and Wellcome Trust [092809/Z/10/Z].

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Susanne Gräslund
    • 1
  • Pavel Savitsky
    • 2
  • Susanne Müller-Knapp
    • 2
    • 3
  1. 1.Structural Genomics Consortium, Department of Biochemistry and BiophysicsKarolinska InstitutetSolnaSweden
  2. 2.Target Discovery Institute and Structural Genomics ConsortiumOxford UniversityOxfordUK
  3. 3.Goethe-University FrankfurtBuchmann Institute for life SciencesFrankfurt am MainGermany

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