Unravelling the fluorescence kinetics of light-harvesting proteins with simulated measurements

• Published in: Biochimica et biophysica acta. Bioenergetics Vol. 1865; no. 1; p. 149004
• Main Authors: Gray, Callum, Kailas, Lekshmi, Adams, Peter G., Duffy, Christopher D.P.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.01.2024
• Subjects: Fluorescence; Fluorescence lifetime; Light-Harvesting Protein Complexes - metabolism; Non-photochemical quenching; Photosynthetic light-harvesting; Photosystem II; Photosystem II Protein Complex - metabolism; Physics; Thylakoids - metabolism; Time-correlated single photon counting; Time-correlated single photon counting; Photosystem II; Fluorescence lifetime; Photosynthetic light-harvesting; Non-photochemical quenching
• ISSN: 0005-2728; 1879-2650
• DOI: 10.1016/j.bbabio.2023.149004

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. •Simulated TCSPC measurements on aggregates of LHCII•Captures underlying physics plus signal detection and data analysis•Allows real TCSPC measurements to yield significant kinetic detail•Quenching in LHCII aggregates results from large sub-population of slow quenchers.•At least two quenching mechanisms needed to explain TCSPC kinetics of LHCII.