Unravelling the fluorescence kinetics of light-harvesting proteins with simulated measurements.
Unravelling the fluorescence kinetics of light-harvesting proteins with simulated measurements.
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DOI：
10.1016/j.bbabio.2023.149004
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发表时间：
2023-07
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影响因子：
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通讯作者：
C. Gray;L. Kailas;Peter G. Adams;Christopher D. P. Duffy
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作者：
C. Gray;L. Kailas;Peter G. Adams;Christopher D. P. Duffy
植物光捕获色素-蛋白质复合物 LHCII 是 PSII 的主要天线亚基，通常（尽管不是普遍）认为在高光条件下的光保护能量耗散中发挥作用，这一过程称为非光化学猝灭 (NPQ)。 LHCII 中能量捕获和耗散的基本机制仍然存在争议。人们针对 NPQ 的基本分子细节提出了各种模型，但它们通常基于对孤立复合物的非常相似的瞬态吸收测量的不同解释。在这里，我们提出了淬灭 LHCII 聚集体的荧光衰减动力学的模拟测量，以确定这种相对简单的测量是否可以区分不同的潜在 NPQ 机制。我们不仅模拟基础物理（激发、能量迁移、猝灭和单线态-单线态湮灭），还模拟信号检测和典型实验数据分析。将其与已发表的荧光衰变动力学进行比较，我们发现：（1）即使在低（无湮灭）激发密度下，不同的拟议猝灭机制也会产生明显不同的荧光动力学，尽管差异程度取决于脉冲宽度。 (2) 测得的衰减动力学与大多数 LHCII 三聚体成为相对较慢的激发猝灭剂一致。非常快的猝灭剂的一小部分产生的动力学与任何观察到的测量结果都不相似。 (3)有必要考虑至少两种不同的猝灭机制，以便准确地再现实验动力学，支持NPQ不是简单的二元开关的观点。
The plant light-harvesting pigment-protein complex LHCII is the major antenna sub-unit of PSII and is generally (though not universally) accepted to play a role in photoprotective energy dissipation under high light conditions, a process known Non-Photochemical Quenching (NPQ). The underlying mechanisms of energy trapping and dissipation within LHCII are still debated. Various models have been proposed for the underlying molecular detail of NPQ, but they are often based on different interpretations of very similar transient absorption measurements of isolated complexes. Here we present a simulated measurement of the fluorescence decay kinetics of quenched LHCII aggregates to determine whether this relatively simple measurement can discriminate between different potential NPQ mechanisms. We simulate not just the underlying physics (excitation, energy migration, quenching and singlet-singlet annihilation) but also the signal detection and typical experimental data analysis. Comparing this to a selection of published fluorescence decay kinetics we find that: (1) Different proposed quenching mechanisms produce noticeably different fluorescence kinetics even at low (annihilation free) excitation density, though the degree of difference is dependent on pulse width. (2) Measured decay kinetics are consistent with most LHCII trimers becoming relatively slow excitation quenchers. A small sub-population of very fast quenchers produces kinetics which do not resemble any observed measurement. (3) It is necessary to consider at least two distinct quenching mechanisms in order to accurately reproduce experimental kinetics, supporting the idea that NPQ is not a simple binary switch.