Association of fluorescent protein pairs and it's significant impact on fluorescence and energy transfer

Dan W Watkins, D. Dafydd Jones*, et al.

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)peer-review

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Abstract


Fluorescent proteins (FPs) are commonly used in pairs to monitor dynamic biomolecular events through changes in proximity via distance dependent processes such as Förster resonance energy transfer (FRET). The impact of FP association is assessed by predicting dimerization sites in silico and stabilizing the dimers by bio‐orthogonal covalent linkages. In each tested case dimerization changes inherent fluorescence, including FRET. GFP homodimers demonstrate synergistic behavior with the dimer being brighter than the sum of the monomers. The homodimer structure reveals the chromophores are close with favorable transition dipole alignments and a highly solvated interface. Heterodimerization (GFP with Venus) results in a complex with ≈87% FRET efficiency, significantly below the 99.7% efficiency predicted. A similar efficiency is observed when the wild‐type FPs are fused to a naturally occurring protein–protein interface system. GFP complexation with mCherry results in loss of mCherry fluorescence. Thus, simple assumptions used when monitoring interactions between proteins via FP FRET may not always hold true, especially under conditions whereby the protein–protein interactions promote FP interaction
Original languageEnglish
Article number2003167
Number of pages11
JournalAdvanced Science
Volume8
Issue number1
Early online date23 Nov 2020
DOIs
Publication statusPublished - 7 Jan 2021

Keywords

  • fluorescence
  • fluorescent proteins
  • Förster resonance energy transfer (FRET)
  • oligomerization
  • protein design

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