Comparison of affinity tags for protein purification

Protein Expr Purif. 2005 May;41(1):98-105. doi: 10.1016/j.pep.2005.01.019.

Abstract

Affinity tags are highly efficient tools for purifying proteins from crude extracts. To facilitate the selection of affinity tags for purification projects, we have compared the efficiency of eight elutable affinity tags to purify proteins from Escherichia coli, yeast, Drosophila, and HeLa extracts. Our results show that the HIS, CBP, CYD (covalent yet dissociable NorpD peptide), Strep II, FLAG, HPC (heavy chain of protein C) peptide tags, and the GST and MBP protein fusion tag systems differ substantially in purity, yield, and cost. We find that the HIS tag provides good yields of tagged protein from inexpensive, high capacity resins but with only moderate purity from E. coli extracts and relatively poor purification from yeast, Drosophila, and HeLa extracts. The CBP tag produced moderate purity protein from E. coli, yeast, and Drosophila extracts, but better purity from HeLa extracts. Epitope-based tags such as FLAG and HPC produced the highest purity protein for all extracts but require expensive, low capacity resin. Our results suggest that the Strep II tag may provide an acceptable compromise of excellent purification with good yields at a moderate cost.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Affinity Labels*
  • Animals
  • Chromatography, Affinity
  • DNA-Binding Proteins / biosynthesis
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / isolation & purification
  • Drosophila
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • HeLa Cells
  • Humans
  • Protein Kinases / biosynthesis
  • Protein Kinases / genetics
  • Protein Kinases / isolation & purification
  • Proteomics
  • Recombinant Fusion Proteins / biosynthesis
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / isolation & purification*
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins / biosynthesis
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / isolation & purification
  • Tetrahydrofolate Dehydrogenase / biosynthesis
  • Tetrahydrofolate Dehydrogenase / genetics
  • Tetrahydrofolate Dehydrogenase / isolation & purification

Substances

  • Affinity Labels
  • DNA-Binding Proteins
  • Recombinant Fusion Proteins
  • Saccharomyces cerevisiae Proteins
  • Tetrahydrofolate Dehydrogenase
  • Protein Kinases