Molecular Mechanisms of Photosynthesis. Robert E. Blankenship
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reciprocal of the quantum requirement) at long wavelengths came to be known as the “red drop.” The interpretation of the red drop is that those chlorophyll molecules that absorb light at the extreme red edge of the absorption band do not do photosynthesis as efficiently as the chlorophylls that absorb light of shorter wavelengths. The long‐wavelength chlorophylls somehow behaved differently. Other measurements of photosynthesis and chlorophyll fluorescence in red algae, organisms that contain an antenna complex known as a phycobilisome, also suggested that the long‐wavelength chlorophylls were somehow inactive in photosynthesis. The red drop result was easily reproduced, but the significance of it was not understood until later.
However, the result of another experiment by Emerson and coworkers was even more bizarre (Emerson et al., 1957). He found that if the ineffective long‐wavelength light was supplemented with shorter‐wavelength light, it suddenly became capable of driving photosynthesis at good rates. A sample of algae was illuminated with red light, and the intensity adjusted to give a particular rate of O2 production, measured as always using a manometer. This light was then turned off, and a second light source, this time the inefficient far‐red light, was directed on the sample. The intensity of this light was adjusted to give a rate of O2 production comparable to that of the red light. This required that the intensity of the far‐red light be increased significantly, as expected from the earlier experiments that had shown its weak effect. The remarkable result was that, when both beams of light were directed on the sample at the same time, the rate of O2 production was greatly increased and was much higher than the sum of the two individual rates! This result came to be known as the enhancement effect, because of the enhancing effect of the short‐wavelength light. Additional experiments by Jack Myers and Stacey French (1960) showed that enhancement worked even when the two beams of light were not present at exactly the same time. These results made no sense in the context of the 1950s understanding of the mechanism of photosynthesis. Several years went by before a reasonable explanation was proposed for these and other puzzling results.
Figure 3.4 Absorption spectrum of chloroplasts (dashed line) and action spectrum for photosynthesis (dotted line). The red drop in the quantum yield of photosynthesis (solid line).
3.11 Antagonistic effects
A final experiment that pointed the way to the existence of two distinct photochemical systems working in series in photosynthetic organisms was carried out by Louis Duysens and coworkers from the Netherlands (Duysens et al., 1961). Duysens was a pioneer in developing sensitive spectrophotometric methods to monitor photosynthetic systems. The experiment was to measure the oxidation–reduction state of cytochrome f in the sample upon illumination using various wavelengths of light. When the cytochrome is reduced, the absorbance spectrum changes, permitting quantitative measurements of its redox state (Fig. 3.5). Duysens found that far‐red light caused the cytochrome to become oxidized, whereas shorter‐wavelength light caused it to become reduced. The two colors of light had opposite, or antagonistic, effects. A particularly clear effect was observed using the red alga Porphyridium cruentum, which has phycobilisome antenna complexes. The effects are easily observed with this organism, because, as we now know, the phycobilisome antenna complex preferentially directs excitations mostly to one of the two photosystems. As is often the case, the choice of the experimental system in which an effect is emphasized was important to the initial understanding of the effect. Subsequent measurements have shown that the effect is, of course, a general one, observed in all oxygen‐evolving photosynthetic organisms.
Figure 3.5 Antagonistic effects on cytochrome oxidation. Irradiation with the light of one color causes the cytochrome to become more oxidized, while irradiation with light of a different color causes it to become more reduced. This experiment was the clearest early evidence for two photochemical systems connected in series in oxygenic photosynthetic organisms.
Source: Duysens et al. (1961)/Springer Nature.
3.12 Early formulations of the Z scheme for photosynthesis
All these experiments (and some others not discussed here) suddenly crystallized into a consistent formulation for photosynthesis about 1960. Robin Hill and Fay Bendall published a short paper in Nature in 1960 outlining the concept of two sequential photochemical systems arranged in tandem, so that the products of one system became the substrates of the other system (Hill and Bendall, 1960). Their formulation was based primarily on the observation that the redox potentials for two cytochromes found in chloroplasts were intermediate between the potentials of both the reductant (H2O) and the oxidant (NADP+) involved in photosynthesis. In order for them to be participants in the light‐driven electron flow, it was necessary to propose two photochemical processes and an energetically downhill intermediate step. Their original scheme is shown in Fig. 3.6. It has come to be known as the Z (for zigzag) scheme of photosynthesis (Govindjee and Björn, 2017).
Figure 3.6 Hill and Bendall's (1960) original formulation of the Z scheme for photosynthesis. Modern Z schemes are plotted with the y‐axis reversed in sign, so that more reducing species are at the top of the diagram and more oxidizing species are at the bottom. The action of light is shown as a vertical arrow in which a relatively highly oxidizing species is converted to a highly reducing one by the action of light.
Source: Hill and Bendall (1960) (p. 137)/Springer Nature.
The slightly later publication by Duysens and coworkers provided the crucial experimental evidence for this proposal, in the form of the antagonistic effects described above. One photochemical reaction, now known as Photosystem II, oxidizes water and reduces cytochrome f, while the other, now known as Photosystem I, oxidizes cytochrome f and reduces NADP+ (Duysens et al., 1961; Duysens, 1989).
The puzzling results of the red drop and enhancement effects are easily explained by this formulation of two photochemical reactions connected in series, if the absorption spectra of the pigments that feed energy to them are not quite the same. The shorter‐wavelength chlorophylls (and the phycobilisome antennas in those organisms that contain them) preferentially drive Photosystem II, whereas the longer‐wavelength pigments preferentially drive Photosystem I. Optimum rates of photosynthesis are observed when both short and long wavelengths are present, as found in the enhancement experiments. The light absorbed by the long‐wavelength pigments does not have enough energy to drive Photosystem II, so the entire system grinds to a halt if only far‐red light is used, thus explaining the red drop phenomenon. In retrospect, it was fortunate that the two photosystems had somewhat different wavelength optima, as the key early experiments demonstrating their existence all relied on preferential excitation of one or the other photosystem by carefully selecting the illumination regime.
The series formulation for oxygen‐evolving photosynthesis has been tested and questioned many times since 1960 and has withstood all challenges. There is now no doubt that this basic framework for photosynthetic electron flow in oxygen‐evolving organisms