Crystal Structure of Phototoxic Orange Fluorescent Proteins With a Tryptophan-Based Chromophore

PLoS One. 2015 Dec 23;10(12):e0145740. doi: 10.1371/journal.pone.0145740. eCollection 2015.

Abstract

Phototoxic fluorescent proteins represent a sparse group of genetically encoded photosensitizers that could be used for precise light-induced inactivation of target proteins, DNA damage, and cell killing. Only two such GFP-based fluorescent proteins (FPs), KillerRed and its monomeric variant SuperNova, were described up to date. Here, we present a crystallographic study of their two orange successors, dimeric KillerOrange and monomeric mKillerOrange, at 1.81 and 1.57 Å resolution, respectively. They are the first orange-emitting protein photosensitizers with a tryptophan-based chromophore (Gln65-Trp66-Gly67). Same as their red progenitors, both orange photosensitizers have a water-filled channel connecting the chromophore to the β-barrel exterior and enabling transport of ROS. In both proteins, Trp66 of the chromophore adopts an unusual trans-cis conformation stabilized by H-bond with the nearby Gln159. This trans-cis conformation along with the water channel was shown to be a key structural feature providing bright orange emission and phototoxicity of both examined orange photosensitizers.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, N.I.H., Intramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Amino Acid Substitution
  • Crystallography, X-Ray
  • Luminescent Proteins / chemistry*
  • Luminescent Proteins / genetics
  • Models, Molecular
  • Molecular Conformation
  • Molecular Sequence Data
  • Mutagenesis
  • Mutation / genetics
  • Photochemical Processes
  • Photosensitizing Agents / chemistry*
  • Protein Conformation
  • Sequence Homology, Amino Acid
  • Tryptophan / chemistry*
  • Tryptophan / genetics

Substances

  • Luminescent Proteins
  • Photosensitizing Agents
  • Tryptophan