Taguchi Masahiko, Sakuraba Shun, Chan Justin, Kono Hidetoshi
Institute for Quantum Life Science, National Institutes for Quantum Science and Technology, Chiba 263-8555, Japan.
Institute of Multidisciplinary Research for Advanced Materials, Tohoku University, Sendai 980-8577, Japan.
ACS Phys Chem Au. 2024 Oct 29;4(6):647-659. doi: 10.1021/acsphyschemau.4c00040. eCollection 2024 Nov 27.
OaPAC is a photoactivated enzyme that forms a homodimer. The two blue-light using flavin (BLUF) photoreceptor domains are connected to the catalytic domains with long coiled-coil C-terminal helices. Upon photoreception, reorganization of the hydrogen bonding network between Tyr6, Gln48, and the chromophore in the BLUF domain and keto-enol tautomerization of Gln48 are thought to occur. However, the quantitative energetics of the photoisomerization reaction and how the BLUF domain's structural change propagates toward the catalytic domain are still unknown. We evaluate the free-energy differences among the dark-state and two different light-state structures by the free-energy perturbation calculations combined with QM/MM free-energy optimizations. Furthermore, we performed long-time MD simulations for the free-energetically optimized dark- and light-state structures to clarify the differences in protein dynamics upon photoisomerization. The free-energy difference between the two optimized light-state structures was estimated at ∼4.7 kcal/mol. The free-energetically optimized light-state structure indicates that the chemically unstable enol tautomer of Gln48 in the light state is stabilized by forming a strong hydrogen bonding network with the chromophore and Tyr6. In addition, the components of free-energy difference between the dark- and light-state structures show that the energy upon photoreception is stored in the environment rather than the internal photoreceived region, suggesting a mechanism to keep the photoactivated signaling state with the chemically unstable enol tautomer of Gln48. In the light state, a fluctuation of Trp90 near the C-terminal helix becomes large, which causes subsequent structural changes in the BLUF core and the C-terminal helix. We also identified residue pairs with significant differences concerning residue-wise contact maps between the dark and light states.
OaPAC是一种形成同型二聚体的光激活酶。两个利用黄素的蓝光（BLUF）光感受器结构域通过长的卷曲螺旋C末端螺旋与催化结构域相连。在光接收时，人们认为BLUF结构域中Tyr6、Gln48和发色团之间的氢键网络会发生重组，并且Gln48会发生酮-烯醇互变异构。然而，光异构化反应的定量能量学以及BLUF结构域的结构变化如何向催化结构域传播仍然未知。我们通过结合QM/MM自由能优化的自由能微扰计算来评估暗态和两种不同光态结构之间的自由能差异。此外，我们对自由能优化后的暗态和光态结构进行了长时间的分子动力学模拟，以阐明光异构化时蛋白质动力学的差异。两种优化后的光态结构之间的自由能差异估计约为4.7千卡/摩尔。自由能优化后的光态结构表明，光态下Gln48化学不稳定的烯醇互变异构体通过与发色团和Tyr6形成强氢键网络而得以稳定。此外，暗态和光态结构之间自由能差异的组成部分表明，光接收时的能量存储在环境中而非内部光接收区域，这表明存在一种机制可以通过Gln48化学不稳定的烯醇互变异构体来维持光激活信号状态。在光态下，C末端螺旋附近的Trp90波动变大，这会导致BLUF核心和C末端螺旋随后发生结构变化。我们还确定了暗态和光态之间在残基接触图方面存在显著差异的残基对。
