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Keywords:

  • fluorescence;
  • FRET (fluorescence resonance energy transfer);
  • proteins;
  • single-molecule studies;
  • time-resolved spectroscopy

Abstract

Transient absorption spectroscopy with sub-100 fs time resolution was performed to investigate the oligomerisation behaviour of eYFP in solution. A single time constant τAD=2.2±0.15 ps is sufficient to describe the time-resolved anisotropy decay up to at least 200 ps. The close contact of two protein barrels is deduced as the exclusive aggregation state in solution. From the final anisotropy r=0.28±0.02, the underlying quaternary structure can be traced back to the somewhat distorted structure of the dimers of wt-GFP. The use of autofluorescent proteins as rulers in Förster resonance energy transfer (FRET) measurements may demand polarisation-sensitive detection of the fluorescence with high time resolution.