Photoinduced electron transfer in singly labeled thiouredopyrenetrisulfonate azurin derivatives

N. Borovok, Alexander Kotlyar*, I. Pecht, L. K. Skov, O. Farver

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


A novel method for the initiation of intramolecular electron transfer reactions in azurin is reported. The method is based on laser photoexcitation of covalently attached thiouredopyrenetrisulfonate (TUPS), the reaction that generates the low potential triplet state of the dye with high quantum efficiency. TUPS derivatives of azurin, singly labeled at specific lysine residues, were prepared and purified to homogeneity by ion exchange HPLC. Transient absorption spectroscopy was used to directly monitor the rates of the electron transfer reaction from the photoexcited triplet state of TUPS to Cu(II) and the back reaction from Cu(I) to the oxidized dye. For all singly labeled derivatives, the rate constants of copper ion reduction were one or two orders of magnitude larger than for its reoxidation, consistent with the larger thermodynamic driving force for the former process. Using 3-D coordinates of the crystal structure of Pseudomonas aeruginosa azurin and molecular structure calculation of the TUPS modified proteins, electron transfer pathways were calculated. Analysis of the results revealed a good correlation between separation distance from donor to Cu ligating atom (His-N or Cys-S) and the observed rate constants of Cu(II) reduction. Copyright (C) 1999 Federation of European Biochemical Societies.

Original languageEnglish
Pages (from-to)277-282
Number of pages6
JournalFEBS Letters
Issue number2
StatePublished - 27 Aug 1999


  • Azurin
  • Electron transfer
  • Pathway calculation
  • Time-resolved spectroscopy
  • Transient kinetics


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