Amino Acids

, Volume 47, Issue 4, pp 729–734

FRET-based analysis of protein-nucleic acid interactions by genetically incorporating a fluorescent amino acid

Original Article

Abstract

Protein–nucleic acid interaction is an important process in many biological phenomena. In this study, a fluorescence resonance energy transfer (FRET)-based protein–DNA binding assay has been developed, in which a fluorescent amino acid is genetically incorporated into a DNA-binding protein. A coumarin-containing amino acid was incorporated into a DNA-binding protein, and the mutant protein specifically produced a FRET signal upon binding to its cognate DNA labeled with a fluorophore. The protein–DNA binding affinity was then measured under equilibrium conditions. This method is advantageous for studying protein-nucleic acid interactions, because it is performed under equilibrium conditions, technically easy, and applicable to any nucleic acid-binding protein.

Keywords

Protein–DNA interaction Unnatural amino acids Fluorescence resonance energy transfer 

Supplementary material

726_2014_1900_MOESM1_ESM.docx (67 kb)
Supplementary material 1 (DOCX 67 kb)

Copyright information

© Springer-Verlag Wien 2014

Authors and Affiliations

  1. 1.Department of ChemistrySogang UniversitySeoulRepublic of Korea

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