Two pyrenylalanines in dihydrofolate reductase form an excimer enabling the study of protein dynamics

Shengxi Chen, Lin Wang, Nour Eddine Fahmi, Stephen J. Benkovic, Sidney M. Hecht

Research output: Contribution to journalArticlepeer-review

28 Scopus citations

Abstract

Because of the lack of sensitivity to small changes in distance by available FRET pairs (a constraint imposed by the dimensions of the enzyme), a DHFR containing two pyrene moieties was prepared to enable the observation of excimer formation. Pyren-1-ylalanine was introduced into DHFR positions 16 and 49 using an in vitro expression system in the presence of pyren-1-ylalanyl- tRNACUA. Excimer formation (λex 342 nm; λem 481 nm) was observed in the modified DHFR, which retained its catalytic competence and was studied under multiple and single turnover conditions. The excimer appeared to follow a protein conformational change after the H transfer involving the relative position and orientation of the pyrene moieties and is likely associated with product dissociation.

Original languageEnglish (US)
Pages (from-to)18883-18885
Number of pages3
JournalJournal of the American Chemical Society
Volume134
Issue number46
DOIs
StatePublished - Nov 21 2012

All Science Journal Classification (ASJC) codes

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

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