The origin of chlorophyll fluorescence In vivo and its quenching by the photosystem II reaction centre

Abstract Isolated chlorophyll a, in contrast to when it is dissolved in organic solvents, shows a lower and variable yield of fluorescence when bound to protein and embedded in the thylakoid membrane of photosynthetic organisms. There are two current theories that attempt to explain the origin of this variable yield of fluorescence, (i) It may be emitted directly from the photosystem II (PSII) antenna system and therefore in competition with photochemical trapping (prompt fluorescence), (ii) It may be derived from a recombination reaction between oxidized P680 and reduced pheophytin within the PS II reaction centre (delayed fluorescence). We have isolated a PS II reaction centre complex that binds only four chlorophyll a molecules and can carry out primary charge separation. The complex contains no plastoquinone and therefore is devoid of the secondary electron acceptor QA. It does, however, contain two pheophytin a molecules, and one of these acts as a primary electron acceptor. The electron donor is P680, which is either a monomeric or dimeric form of chlorophyll a. The isolated PS II reaction centre fluoresces at room temperature with a maximum at 683 nm, and the intensity of this emission is almost totally quenched when reduced pheophytin (bright light plus sodium dithionite) or oxidized P680 (bright light plus silicomolybdate) is photoaccumulated. The photo-induced quenching of chlorophyll fluorescence when sodium dithionite is present is also observed in intact PS II preparations containing plastoquinone QA. In the latter case QA is chemically reduced in the dark by dithionite. Bearing in mind the above two postulates for the origin of variable chlorophyll fluorescence it has been possible to investigate the relative quantum yields for the photoproduction of the P680Pheo- state either in the absence (with isolated PS II reaction centres) or presence (with PSII-enriched membranes) of reduced QA. It has been shown that in the absence of Q-A the quantum efficiency for production of the P680Pheo- is several orders of magnitude greater than when Q-A is present. This difference probably partly reflects the coulombic restraints on primary charge separation when QA is reduced and would suggest that under these conditions the PS II reaction centre is a less efficient trap. Such a conclusion is therefore consistent with postulate (i) that the increase inyield of chlorophyll fluorescence as QA becomes reduced is not due to a back reaction between P+680 and Pheo- but rather to a decrease in competition between emission and trapping. The results do emphasize however, that the P680Pheo- and P+680Pheo states are quenchers of chlorophyll fluorescence. In addition to the above, it has been noted that at 77 K fluorescence from the isolated PS II reaction centre reaches a maximum at 685 nm and does not have a peak at 695 nm. This observation appears to invalidate the postulate that the 695 nm emission is from the pheophytin of the PS II reaction centre.