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

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-coumari...

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Bibliographic Details
Published in:Biophysical journal 2017-05, Vol.112 (10), p.2109-2116
Main Authors: Oktaviani, Nur Alia, Pool, Trijntje J., Yoshimura, Yuichi, Kamikubo, Hironari, Scheek, Ruud M., Kataoka, Mikio, Mulder, Frans A.A.
Format: Article
Language:English
Subjects:
Aspartic Acid - chemistry
Aspartic Acid - metabolism
Bacteria
Bacterial Proteins - chemistry
Bacterial Proteins - metabolism
Coumaric acid
Experiments
Glutamic Acid - chemistry
Glutamic Acid - metabolism
Halorhodospira halophila
Hydrogen Bonding
Hydrogen-Ion Concentration
Kinetics
Light effects
Nuclear Magnetic Resonance, Biomolecular
Optical properties
pH effects
Photoactive yellow protein
Photoreceptors, Microbial - chemistry
Photoreceptors, Microbial - metabolism
Proteins
Protonation
Protons
Recovery
Residues
Yellow protein
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Summary: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.
ISSN:0006-3495
1542-0086
DOI:10.1016/j.bpj.2017.04.008