Structure-based engineering of environmentally sensitive fluorophores for monitoring protein-protein interactions

Protein Eng. 1998 Sep;11(9):819-23. doi: 10.1093/protein/11.9.819.

Abstract

Single, extrinsic, environmentally sensitive fluorophores can be used to quantitate formation of protein-protein complexes. These can be prepared semi-synthetically by covalent coupling to single cysteine mutations introduced at positions where the fluorophore is predicted to respond to formation of the complex without adversely affecting the interaction. The three-dimensional structure of a protein-protein interface can be used to select such locations by identifying residues that are located at the edge of a buried interfacial region, and are in partial steric contact with both partners as indicated by a change in their static solvent-accessible surface area upon complex formation. Using this design approach, cysteine mutations were introduced into the B1 domain of protein G, which successfully monitor complex formation with minimal interference. Such constructs have great utility in the analysis of solution properties of interface mutants.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • DNA Primers
  • Fluorescent Dyes / chemical synthesis*
  • Fluorescent Dyes / chemistry
  • Models, Molecular
  • Molecular Structure
  • Nerve Tissue Proteins / metabolism*
  • Protein Binding

Substances

  • DNA Primers
  • Fluorescent Dyes
  • G-substrate
  • Nerve Tissue Proteins