Evidence for a cysteine-mediated mechanism of excitation energy regulation in a photosynthetic antenna complex

Abstract

Light-harvesting antenna complexes not only aid in the capture of solar energy for photosynthesis, but regulate the quantity of transferred energy as well. Light-harvesting regulation is important for protecting reaction center complexes from overexcitation, generation of reactive oxygen species, and metabolic overload. Usually, this regulation is controlled by the association of lightharvesting antennas with accessory quenchers such as carotenoids. One antenna complex, the Fenna-Matthews-Olson (FMO) antenna protein from green sulfur bacteria, completely lacks carotenoids and other known accessory quenchers. Nonetheless, the FMO protein is able to quench energy transfer in aerobic conditions effectively, indicating a previously unidentified type of regulatory mechanism. Through de novo sequencing MS, chemical modification, and mutagenesis, we have pinpointed the source of the quenching action to cysteine residues (Cys49 and Cys353) situated near two lowenergy bacteriochlorophylls in the FMO protein from Chlorobaculum tepidum. Removal of these cysteines (particularly removal of the completely conserved Cys353) through N-ethylmaleimide modification or mutagenesis to alanine abolishes the aerobic quenching effect. Electrochemical analysis and electron paramagnetic resonance spectra suggest that in aerobic conditions the cysteine thiols are converted to thiyl radicals which then are capable of quenching bacteriochlorophyll excited states through electron transfer photochemistry. This simple mechanism has implications for the design of bio-inspired light-harvesting antennas and the redesign of natural photosynthetic systems.photosynthesis | Fenna-Matthews-Olson protein | excitation quenching | thiyl radical | light-harvesting P hotosynthesis can be performed in the presence of oxygen (oxygenic photosynthesis, in which water is the electron source) or in its absence (anoxygenic photosynthesis, in which other reduced species such as sulfide are the electron sources) (1-3). Many bacteria performing anoxygenic photosynthesis contain type I reaction center (RC) protein complexes that use Fe-S clusters to transfer electrons to ferredoxin or related molecules after photoinduced charge separation (4-6). These Fe-S clusters are easily damaged by molecular oxygen. Therefore, anoxygenic phototrophs must use a pathway either to remove oxygen or to reduce the rate of photosynthesis whenever oxygen is encountered (1, 5).Phototrophic members of the bacterial phylum Chlorobi (green sulfur bacteria, GSBs) are anoxygenic phototrophs that contain such a type I RC. They also contain two peripheral antenna complexes that aid in regulating light absorption and energy transfer: the chlorosome and the homotrimeric Fenna-Matthews-Olson (FMO) protein complex. When oxygen is encountered, the bacteria are able to decrease the output of photosynthesis. This process involves creating trapping sites in the antenna complexes where de-excitation processes outcompete the rate of energy transfer to the RC, preventing RC damage. This process is well un...

Physiological Studies of Chlorobiaceae Suggest that Bacillithiol Derivatives Are the Most Widespread Thiols in Bacteria

“… Chlorobaculum tepidum has previously been suggested to synthesize glutathione ( 44 ), and the Chlorobium limicola genome contains two genes, Clim_1148 and Clim_1149, whose recombinant products convert histidine to trimethylhistidine by a methyltransferase (Clim_1148, EanA) and trimethylhistidine to ergothioneine by a sulfur transferase reaction (Clim_1149, EanB) in vitro ( 26 ). The ergothioneine biosynthetic pathway was thought to be restricted to aerobic organisms, because one of the enzymes in the pathway, EgtB, required molecular oxygen ( 26 ).…”

“…Another possible function in the Chlorobiaceae is that N-Me-BSH is the in vivo reductant for the recently described mechanism for regulating excitation energy transfer in the FmoA protein ( 44 ). Cysteine localized thiyl radicals in FmoA are proposed to interact with excited bacteriochlorophyll a to prevent excitation energy transfer to the reaction center under unfavorable conditions.…”

“…Cysteine localized thiyl radicals in FmoA are proposed to interact with excited bacteriochlorophyll a to prevent excitation energy transfer to the reaction center under unfavorable conditions. In vitro , dithionite, dithiothreitol, sulfide, glutathione, and TCEP were capable of regulating FmoA energy transfer, and GSH was proposed as the in vivo mediator ( 44 ). The results presented here indicate that GSH is not a good candidate for this function, as no Chlorobiaceae genomes encode GSH biosynthesis ( Fig.…”

N-methyl-bacillithiol, a new metabolite discovered in theChlorobiaceae, indicates that bacillithiol and derivatives are widely phylogenetically distributed

“…Another possible function in the Chlorobiaceae is that N-Me-BSH is the in vivo reductant for the recently described excitation energy regulating mechanism in the FmoA protein (38). Cysteine centered thiyl radicals in FmoA are proposed to interact with excited bacteriochlorophyll a to prevent excitation energy transfer to the reaction center under unfavorable conditions.…”

“…Cysteine centered thiyl radicals in FmoA are proposed to interact with excited bacteriochlorophyll a to prevent excitation energy transfer to the reaction center under unfavorable conditions. In vitro , dithionite, dithiothreitol, sulfide, glutathione and TCEP were capable of capable of regulating FmoA energy transfer, and GSH was proposed as the in vivo mediator (38). The results presented here indicate that GSH is not a good candidate as no Chlorobiaceae genomes encode GSH biosynthesis (Fig.…”