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

TY - JOUR

T1 - Active-Site pKa Determination for Photoactive Yellow Protein Rationalizes Slow Ground-State Recovery

AU - Oktaviani, Nur Alia

AU - Pool, Trijntje J

AU - Yoshimura, Yuichi

AU - Kamikubo, Hironari

AU - Scheek, Ruud M

AU - Kataoka, Mikio

AU - Mulder, Frans A.A.

N1 - Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

PY - 2017/5/23

Y1 - 2017/5/23

N2 - 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.

AB - 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.

KW - Journal Article

UR - http://www.scopus.com/inward/record.url?scp=85019717154&partnerID=8YFLogxK

U2 - 10.1016/j.bpj.2017.04.008

DO - 10.1016/j.bpj.2017.04.008

M3 - Journal article

C2 - 28538148

SN - 0006-3495

VL - 112

SP - 2109

EP - 2116

JO - Biophysical Journal

JF - Biophysical Journal

IS - 10

ER -