Zeaxanthin Radical Cation Formation in Minor Light-harvesting Complexes of Higher Plant Antenna

Abstract

Previous work on intact thylakoid membranes showed that transient formation of a zeaxanthin radical cation was correlated with regulation of photosynthetic light-harvesting via energy-dependent quenching. A molecular mechanism for such quenching was proposed to involve charge transfer within a chlorophyll-zeaxanthin heterodimer. Using near infrared (880 -1100 nm) transient absorption spectroscopy, we demonstrate that carotenoid (mainly zeaxanthin) radical cation generation occurs solely in isolated minor light-harvesting complexes that bind zeaxanthin, consistent with the engagement of charge transfer quenching therein. We estimated that less than 0.5% of the isolated minor complexes undergo charge transfer quenching in vitro, whereas the fraction of minor complexes estimated to be engaged in charge transfer quenching in isolated thylakoids was more than 80 times higher. We conclude that minor complexes which bind zeaxanthin are sites of charge transfer quenching in vivo and that they can assume Non-quenching and Quenching conformations, the equilibrium LHC(N) % LHC(Q) of which is modulated by the transthylakoid pH gradient, the PsbS protein, and protein-protein interactions.Higher plant photosynthesis is initiated by absorption of light in pigment-binding (antenna) proteins that transfer absorbed solar energy to the reaction centers of photosystems (PS) 3 II and I where energy conversion begins (1). The PSIIassociated light-harvesting complexes (LHCs) bind chlorophylls and carotenoids that are involved in both the harvesting and transfer of energy to the reaction center, and the harmless dissipation of excitation energy in excess of photosynthetic capacity (2). Thus, the PSII LHCs are critical branch-points for energy partitioning during photosynthesis. The peripheral antenna consists of trimeric complexes composed of LHCII proteins, the major LHC of higher plant antennae. In between the peripheral LHCII and the reaction center there are three minor LHCs referred to as CP24, CP26, and CP29 (1).Dissipation of excess light energy during photosynthesis involves several photoprotective mechanisms, which are collectively referred to as non-photochemical quenching (NPQ) (2, 3). The predominant component of NPQ is referred to as energy-dependent quenching, or qE, and it is rapidly reversible and correlated with zeaxanthin (Z) formation (4). Mutants of Arabidopsis thaliana have been instrumental in confirming the involvement of Z (5) and identifying a role for PsbS in qE (6). The npq4 mutant lacks a functional PsbS protein and exhibits very little qE (6). The PsbS protein has been subsequently proposed to be involved in controlling qE by sensing thylakoid lumen pH (7). The xanthophyll cycle consists of the enzymatic and reversible conversion of the thylakoid-associated pigment violaxanthin (V) to antheraxanthin (A) and Z (8). Very little qE is exhibited in the A. thaliana mutant referred to as npq1 which is impaired in its ability to convert V to Z as a result of a lesion in the gene encoding the thylakoid lumen-loca...

Snapshot Transient Absorption Spectroscopy of Carotenoid Radical Cations in High-Light-Acclimating Thylakoid Membranes

“…We suggest that this is due to trimeric LHCII migration away from the photosystem II (PSII) supercomplex, which is proposed to occur within 5 min of high light exposure. 15,16 If, as suggested previously, 11,12,17 CT quenching occurs in the monomeric LHCII, this reduces the amount of excitation energy funneled to the CT quenching site. It is noteworthy that the Zea •+ TA signal slowly decreased in dark periods despite the near-constant Zea concentration, indicative of asymmetric induction-relaxation of the CT quenching mechanism.…”

“…6 However, these thylakoids had been high-light-acclimated for over 30 min before measurement, which does not indicate whether CT quenching is activated within the first few minutes of high light exposure, the time scale of qE activation. Zea •+ has also been observed in isolated minor (monomeric) light-harvesting complexes containing Zea, 11,12 but these protein conditions may not be indicative of in vivo behavior, and once again, do not give information about when CT quenching turns on during light acclimation. In a recent study, Dall'Osto and coworkers concluded that the trimeric light-harvesting complex II (LHCII) is the location of a more slowly activated (several minutes) quenching mechanism that does not involve formation of Zea •+ in vivo.…”

From ecophysiology to phenomics: some implications of photoprotection and shade–sun acclimation in situ for dynamics of thylakoids in vitro

“…The fact that Chl-Z charge transfer quenching predominantly occurs in minor antenna complexes (Lhcb4, 5 and 6) and not in LHCII [53] is in line with the higher Chl fluorescence yield measured in Lhcb5 reconstituted with Lx but not in Lhcb1 and trimeric LHCII [32]. Interestingly, recombinant Arabidopsis Lhcb5 exhibits carotenoid radical cations in both L1 (with L) and L2 (with Z) sites although binding of Z in L2 seems to be a prerequisite for Chl-L charge transfer quenching in L1 [54].…”

On the origin of a slowly reversible fluorescence decay component in the Arabidopsis npq4 mutant

“…As NPQ depends on the antenna proteins [14,22,23,36], we evaluated the capacity of npq4 mutants, devoid of specific LHC gene products, to modulate qM. In mutants devoid of both Lhcb5 and Lhcb6 subunits ( figure 3c and table 2), the amplitude and relaxation of NPQ were essentially the same as observed in npq4 mutants.…”