Active-Site pKa Determination for Photoactive Yellow Protein Rationalizes Slow Ground-State Recovery

Publication: Research - peer-reviewJournal article

DOI

  • Nur Alia Oktaviani
    Nur Alia OktavianiGroningen Biomolecular Sciences and Biotechnology Institute, University of GroningenNetherlands
  • Trijntje J Pool
    Trijntje J PoolGroningen Biomolecular Sciences and Biotechnology Institute, University of GroningenNetherlands
  • Yuichi Yoshimura
    Yuichi Yoshimura
  • Hironari Kamikubo
    Hironari KamikuboNara Institute of Science and TechnologyJapan
  • Ruud M Scheek
    Ruud M ScheekGroningen Biomolecular Sciences and Biotechnology Institute, University of GroningenNetherlands
  • Mikio Kataoka
    Mikio KataokaNara Institute of Science and TechnologyJapan
  • Frans A.A. Mulder

The ability to avoid blue-light radiation is crucial for bacteria to survive. In Halorhodospira halophila, the putative receptor for this response is known as photoactive yellow protein (PYP). Its response to blue light is mediated by changes in the optical properties of the chromophore para-coumaric acid (pCA) in the protein active site. PYP displays photocycle kinetics with a strong pH dependence for ground-state recovery, which has remained enigmatic. To resolve this problem, a comprehensive pKa determination of the active-site residues of PYP is required. Herein, we show that Glu-46 stays protonated from pH 3.4 to pH 11.4 in the ground (pG) state. This conclusion is supported by the observed hydrogen-bonded protons between Glu-46 and pCA and Tyr-42 and pCA, which are persistent over the entire pH range. Our experimental results show that none of the active-site residues of PYP undergo pH-induced changes in the pG state. Ineluctably, the pH dependence of pG recovery is linked to conformational change that is dependent upon the population of the relevant protonation state of Glu-46 and the pCA chromophore in the excited state, collaterally explaining why pG recovery is slow.

Original languageEnglish
JournalBiophysical Journal
Volume112
Issue number10
Pages (from-to)2109-2116
Number of pages8
ISSN0006-3495
DOIs
StatePublished - 2017

    Keywords

  • Journal Article

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