Tryptophan-based fluorophores for studying protein conformational changes

Poulami Talukder, Shengxi Chen, C. Tony Liu, Edwin A. Baldwin, Stephen J. Benkovic, Sidney Hecht

Research output: Contribution to journalArticlepeer-review

26 Scopus citations


With the continuing interest in deciphering the interplay between protein function and conformational changes, small fluorescence probes will be especially useful for tracking changes in the crowded protein interior space. Presently, we describe the potential utility of six unnatural amino acid fluorescence donors structurally related to tryptophan and show how they can be efficiently incorporated into a protein as fluorescence probes. We also examine the various photophysical properties of the new Trp analogues, which are significantly redshifted in their fluorescence spectra relative to tryptophan. In general, the Trp analogues were well tolerated when inserted into Escherichia coli DHFR, and did not perturb enzyme activity, although substitution for Trp22 did result in a diminution in DHFR activity. Further, it was demonstrated that D and E at position 37 formed efficient FRET pairs with acridon-2-ylalanine (Acd) at position 17. The same was also true for a DHFR construct containing E at position 79 and Acd at position 17. Together, these findings demonstrate that these tryptophan analogues can be introduced into DHFR with minimal disruption of function, and that they can be employed for the selective study of targeted conformational changes in proteins, even in the presence of unmodified tryptophans.

Original languageEnglish (US)
Pages (from-to)5924-5934
Number of pages11
JournalBioorganic and Medicinal Chemistry
Issue number21
StatePublished - Nov 1 2014


  • Amino acid
  • FRET
  • Fluorescence
  • Protein biosynthesis
  • Tryptophan analogues

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Pharmaceutical Science
  • Drug Discovery
  • Clinical Biochemistry
  • Organic Chemistry


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