Annihilation of Excess Excitations along Phycocyanin Rods Precedes Downhill Flow to Allophycocyanin Cores in the Phycobilisome of Synechococcus elongatus PCC 7942

Abstract

Cyanobacterial phycobilisome complexes absorb visible sunlight and funnel photogenerated excitons to the photosystems where charge separation occurs. In the phycobilisome complex of Synechococcus elongatus PCC 7942, phycocyanin protein rods that absorb bluer wavelengths are assembled on allophycocyanin cores that absorb redder wavelengths. This arrangement creates a natural energy gradient toward the reaction centers of the photosystems. Here, we employ broadband pump–probe spectroscopy to observe the fate of excess excitations in the phycobilisome complex of this organism. We show that excess excitons are quenched through exciton–exciton annihilation along the phycocyanin rods prior to transfer to the allophycocyanin cores. Our observations are especially relevant in comparison to other antenna proteins, where exciton annihilation primarily occurs in the lowest-energy chlorophylls. The observed effect could play a limited photoprotective role in physiological light fluences. The exciton decay dynamics is faster in the intact phycobilisome than in isolated C-phycocyanin trimers studied in earlier work, confirming that this effect is an emergent property of the complex assembly. Using the obtained annihilation data, we calculate exciton hopping times of 2.2–6.4 ps in the phycocyanin rods. This value agrees with earlier FRET calculations of exciton hopping times along phycocyanin hexamers by Sauer and Scheer.

Phycobilisome’s Exciton Transfer Efficiency Relies on an Energetic Funnel Driven by Chromophore–Linker Protein Interactions

“…Our obtained dynamics in the all-parallel data acquisition sequence match earlier reports and compartmental models yielding a sub-5 ps component, an ∼50 ps component and an ∼1 ns component. Similar dynamics have been observed in many studies ,− ,, by fitting transient absorption and time-resolved emission decay traces. Our cross-peak specific spectra show distinctly different dynamics from these reports and our own all-parallel 2DES because the pulse sequence selects the cross-peak signal, or the stimulated emission from energy transfer, while suppressing the highly wavelength-dependent dynamics of the other three signatures.…”

“…Two-dimensional spectra of the phycobilisome obtained with identically linearly polarized pulses at 0.2, 2, 20, and 200 ps (Figure a) show dynamics in agreement with past measurements and significant spectral congestion. − The elongated positive feature along the diagonal from 590 to 670 nm in these spectra at early times corresponds to ground-state bleach and stimulated emission of all chromophores. Phycocyanobilin, the only bilin type in the Synechocystis sp.…”

“…Based on the Förster calculations of Sauer and Scheer, energy flow along rods of phycocyanin hexamers should occur primarily through intertrimer exciton hopping between adjacent β 84 1 and β 84 4 rod chromophores. This hop occurs with a FRET rate of 1/(2.5 ps). , To rationalize the fast downhill energy transfer rate, we initially used a random hopping model of excitons along four isoenergetic β 84 sites with a wall on one end and an allophycocyanin sink on the other. Four sites are used based on the negative staining EM images of Gao and co-workers and other TEM and theoretical studies of phycocyanin assembly and rod-length as functions of light intensity used for cellular growth, which also suggest that two hexamers or four trimers are most commonly found in intact phycobilisome structures. , Considering the allophycocyanin as a sink is not easily justified because many studies suggest that the bottleneck for excitation transfer in the phycobilisome is from the rod to the core because of the large interchromophore distance. , However, even assuming an allophycocyanin sink, we retrieve only a mean phycocyanin to allophycocyanin transfer time of 16 ps (see the Supporting Information), which does not agree with our measurements.…”

“…This hop occurs with a FRET rate of 1/(2.5 ps). 18,33 To rationalize the fast downhill energy transfer rate, we initially used a random hopping model of excitons along four isoenergetic β 84 sites with a wall on one end and an allophycocyanin sink on the other. Four sites are used based on the negative staining EM images of Gao and co-workers 9 and other TEM and theoretical studies of phycocyanin assembly and rod-length as functions of light intensity used for cellular growth, which also suggest that two hexamers or four trimers are most commonly found in intact phycobilisome structures.…”

“…In coherent time-resolved spectra, the phycobilisome shows a broad photoinduced absorption (PIA) feature suggested to arise from an electrochromic shift coupling 21,37,38 of excited-and ground-state chromophores, which masks the red-most absorption and complete emission profile. 18,21,39 Previous time-resolved studies have suggested that excitations created in the rods travel downhill to the higher-energy ApcA and ApcB (hereafter called APC 660 for their emission maxima) core proteins on the 30−50 ps timescale and from APC 660 to the core terminal emitters bound to ApcD, ApcE, and ApcF (hereafter called APC 680 ) proteins, on the 100 ps timescale before fluorescing with ca. 1−2 ns lifetime in detached complexes.…”