Hetero-oligomeric tagging diminishes non-specific aggregation of target proteins fused with Anthozoa fluorescent proteins

Biochem J. 2003 Apr 1;371(Pt 1):109-14. doi: 10.1042/BJ20021796.

Abstract

The tendency for tetramerization is the main disadvantage in the green fluorescent protein homologues from Anthozoa species. We report a universal method called hetero-oligomeric tagging, which diminishes troublesome consequences of tetramerization of Anthozoa-derived fluorescent proteins (FP) in intracellular protein labelling. This approach is based on the co-expression of the FP-tagged protein of interest together with an excess of free non-fluorescent FP mutant. The resulting FP heterotetramers contain only a single target polypeptide and, therefore, can be considered pseudo-monomeric. Feasibility of the method has been demonstrated with a red FP fused with cytoplasmic beta-actin or tubulin-binding protein Tau34. In addition, heterotetramers appeared to be a unique model for biophysical characterization of Anthozoa FPs in pseudo-monomeric state.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actins / genetics
  • Actins / metabolism
  • Animals
  • Anthozoa / chemistry*
  • Biochemistry / methods*
  • CHO Cells
  • Cricetinae
  • Gene Expression Regulation
  • Genes
  • Green Fluorescent Proteins
  • Luminescent Proteins / chemistry
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism*
  • Microtubule-Associated Proteins / genetics
  • Microtubule-Associated Proteins / metabolism
  • Mutation
  • Protein Binding
  • Protein Conformation
  • Protein Engineering / methods*
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism*

Substances

  • Actins
  • Luminescent Proteins
  • Microtubule-Associated Proteins
  • Recombinant Fusion Proteins
  • red fluorescent protein
  • Green Fluorescent Proteins