Sustained photoprotection involves enhanced fluorescence intermittency in a subpopulation of LHCII

Photoprotection against excess energy is essential for the survival of photosynthetic organisms under adverse conditions. In plants, excess energy can be dissipated as heat through non-photochemical quenching (NPQ) of chlorophyll fluorescence, involving the trimeric light-harvesting complex II (LHCII), the major antenna of photosystem II. How NPQ affects antenna proteins remains debated, especially as most studies focus on short-lived components artificially induced in vitro. Here, we characterize the effects of qH, a long-lived NPQ component, on the fluorescence properties of natively quenched LHCII. Single-molecule fluorescence measurements, combined with biochemical and biophysical ensemble approaches, reveal a larger and more quenched subpopulation of LHCII trimers exhibiting fluorescence intermittency in samples with qH compared to those without. This behavior is linked to a small conformational change that stabilizes a quenched state, enhancing photoprotection at the antenna level. These findings provide new insights into sustained NPQ and its role in regulating energy dissipation under natural light conditions.