TALKING POINT The origin and evolution of oxygenic photosynthesis

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4 Hijikata, M. et al. (1991) Proc. Natl. Acad. Sci. U. S. A. 88, 5547–5551 5 Hijikata, M. et al. (1993) J. Virol. 67, 4665–4675 6 Grakoui, A. et al. (1993) Proc. Natl. Acad. Sci. U. S. A. 90, 10583–10587 7 Grakoui, A. et al. (1993) J. Virol. 67, 2832–2843 8 Houghton, M. (1996) in Fields Virology (Vol. 1) (Fields, B. N., Knipe, D. M. and Howley, P. M.,

eds), pp. 1036–1041, Lippincott-Raven 9 Voss, T. et al. (1995) Protein Sci. 4, 2526–2531 10 Sommergruber, W. et al. (1994) Virology 204, 815–818 11 Pieroni, L. et al. (1997) J. Virol. 71, 6373–6380 12 Vallee, B. L. and Auld, D. S. (1990) Biochemistry 29, 5647–5659 13 Springman, E. R. et al. (1990) Proc. Natl. Acad. Sci. U. S. A. 87, 364–368

14 Reed, K. E. et al. (1995) J. Virol. 69, 4127–4136 15 Lipscomb, W. N. and Strater, N. (1996) Chem. Rev. 96, 2375–2433 16 Rawlings, N. D. and Barrett, A. J. (1994) Methods Enzymol. 244, 461–486 17 Rawlings, N. D. and Barrett, A. J. (1997) in Proteolysis in Cell Functions (Hopsuhavu, V. K., Jarvinen, M. and Kirschke, H., eds), pp. 13–21, IOS Press

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The origin and evolution of oxygenic photosynthesis

P870*/P870+ P680*/P680+

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Robert E. Blankenship and Hyman Hartman

THE ADVENT of oxygen-evolving photosynthesis is one of the central events in the development of life on Earth. Before the evolution of this metabolic capability, the atmosphere of the early Earth was largely anaerobic1,2. The development of advanced eukaryotic life forms did not take place until the free oxygen in the atmosphere rose to a sufficient level. While significant questions remain over when oxygenic photosynthesis began3–5, no other known process, either biogenic or non-biogenic, is capable of producing the large quantities of molecular oxygen that demonstrably changed the course of life on Earth. Understanding the evolutionary origin of this metabolic process is therefore of considerable importance. All known oxygen-evolving photosynthetic organisms contain two photosystems linked in series6. Water oxidation is carried out by photosystem II and ferredoxin reduction is mediated by photosystem I. Several groups of anoxygenic (non-oxygen-evolving) photosynthetic R. E. Blankenship is in the Department of Chemistry and Biochemistry, Center for the Study of Early Events in Photosynthesis, Arizona State University, Tempe, AZ 85287-1604, USA. H. Hartman is at the Institute for Advanced Studies in Biology, 880 Spruce St., Berkeley, CA 94707, USA.

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bacteria contain reaction center complexes that show clear evolutionary relatedness to one or other of the two photosystems found in oxygenic organisms and are thought to be more closely related to the earliest photosynthetic organisms7–10. However, no native organisms containing only a single photosystem are known that can oxidize water. Also, all known oxygen-evolving organisms contain the photosynthetic pigment chlorophyll a, while anoxygenic photosynthetic bacteria contain one of several bacteriochlorophylls (a, b or g) as reaction center pigments, all of which absorb longer wavelength, and therefore lower energy, light than does chlorophyll a. The major consequence of this is that the excited state energy and therefore the redox potential span that can be generated by bacteriochlorophyllcontaining organisms is significantly less than in chlorophyll-containing organisms (see Fig. 1). Oxygenic photosynthesis is found in cyanobacteria and related prokaryotes, and in a variety of eukaryotic organisms whose chloroplasts were formed by endosymbiosis of a simpler photosynthetic organism11. The mechanism of oxygen production in all known oxygenic photosynthetic organisms appears to be very similar, and involves charge separation

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The evolutionary developments that led to the ability of photosynthetic organisms to oxidize water to molecular oxygen are discussed. Two major changes from a more primitive non-oxygen-evolving reaction center are required: a charge-accumulating system and a reaction center pigment with a greater oxidizing potential. Intermediate stages are proposed in which hydrogen peroxide was oxidized by the reaction center, and an intermediate pigment, similar to chlorophyll d, was present.

Fe2+/Fe(OH)3 H2O2/O2

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H2O/O2 1.0

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Figure 1 Midpoint reduction potentials at pH 7 and 25⬚C (Em⬘) of various redox couples relevant to photosynthetic electron transfer. All potentials shown are relative to the normal hydrogen electrode and are written as reduced/oxidized forms of the redox couple. The reaction depicted is: oxidized + e – → reduced. Hydrogen ions are not shown. The vertical arrows on the right indicate the photon excitation in the purple bacterial reaction center (P870 → P870*) and the photosystem II reaction center (P680 → P680*). The length of these arrows indicates the excitation energy and also the redox span between the ground and excited state redox potentials27. The redox reactions in the text (Eqns 1 and 2) are indicated as oxidations, although, by convention, the midpoint potentials are given for the reduction reaction. The reaction center potentials are taken from Ref. 27, the Fe2+/Fe(OH)3 potential from Ref. 26 and the H2O and H2O2 potentials from Ref. 34.

Copyright © 1998, Elsevier Science Ltd. All rights reserved. 0968 – 0004/98/$19.00

PII: S0968-0004(98)01186-4

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TIBS 23 – MARCH 1998 by a chlorophyll a-containing photosynthetic reaction center and charge accumulation by a Mn–protein complex prior to the actual conversion of water into molecular oxygen12–15. Water is a very stable compound, and its oxidation to molecular oxygen requires a powerful oxidizing agent (Eqn 1): 2H2O → O2 ⫹ 4H⫹ ⫹ 4e ⫺

(1)

The midpoint reduction potential for this process is E m⬘ = ⫹0.82 V (Fig. 1); thus an oxidant with redox potential greater than 0.82 V is needed to decompose water into molecular oxygen. Two major evolutionary developments are required before water oxidation can take place: the development of an oxidant with a sufficiently high redox potential and the ability to collect and store oxidizing equivalents, formed by the photochemical events in the reaction center, that transfer at most one electron per photon absorbed. Neither of these properties is found in any of the known anoxygenic photosynthetic reaction centers, nor are any intermediate forms known. The midpoint redox potential of P680, the reaction center photoactive chlorophyll of photosystem II, is greater than 1 V (Refs 12, 13), fully half a volt above the bacteriochlorophyll-containing reaction centers found in anoxygenic bacteria16. The simultaneous evolutionary development of both the high redox potential species needed to oxidize water and the charge accumulation system needed to carry out the four-electron chemistry of water splitting (Eqn 1) would seem impossible, as each characteristic is almost certainly the result of several independent molecular changes in the reaction center proteins. What is needed is a series of transitional forms, in which one of these capabilities arises first in a simplified, but still functional, form and the other characteristic then arises later. Olson17 proposed a series of nitrogen compounds of increasing redox potentials as possible transitional electron donors to a reaction center that gradually increased its redox potential as the more easily oxidized species were depleted. However, only one of these compounds is able to be oxidized by any known photosynthetic reaction center, and this proposal also does not provide a plausible transition to the present Mn-containing system in which the electron donor is water. Here, we propose an evolutionary scenario that provides for functional intermediate forms that both benefited the organisms that contained them and link

to the current system. The first proposal is that hydrogen peroxide may have been a transitional electron donor on the early Earth and that the current oxygenevolving complex may be structurally related to Mn-containing catalase enzymes. The second proposal is that a pigment related to chlorophyll d may have been an intermediate pigment between bacteriochlorophyll-containing reaction centers and chlorophyll a-containing reaction centers.

Hydrogen peroxide as a transitional electron donor Hydrogen peroxide is capable of being both an oxidant and a reductant. The oxidation of hydrogen peroxide to oxygen can be carried out by a modestly oxidizing species (Em⬘ = 0.27 V), which is fully within the oxidative capabilities of reaction centers from existing anoxygenic photosynthetic bacteria (Fig. 1). H2O2 → 2H⫹ ⫹ 2e ⫺ ⫹ O2

(2)

A bacteriochlorophyll-containing reaction center is thermodynamically capable of oxidizing hydrogen peroxide to molecular oxygen. This oxidation is similar to part of the mechanism of catalase enzymes. The reaction cycle of catalase involves both oxidation and reduction of hydrogen peroxide18. Mn catalase enzymes are known that have a binuclear metal center that is structurally similar to half of the proposed geometry of the tetranuclear Mn center that makes up the photosynthetic oxygen-evolving complex18,19 (Fig. 2). This structural similarity has been previously noted18–20. Under certain conditions, the photosynthetic Mn complex can act as a catalase21,22. The possibility of hydrogen peroxide serving as a transitional electron donor has also been suggested previously20,22,23. An association of an anoxygenic photosynthetic reaction center and a Mn catalase enzyme might produce a primitive system that evolved oxygen, using hydrogen peroxide as an electron donor. Subsequent developments could convert the binuclear Mn site to a tetranuclear site capable of accumulating up to four oxidizing equivalents. The gene sequence for the Mn catalase from Lactobacillus plantarum has recently been published24. There are no obvious sequence similarities to the loop regions of the D1 protein, which is thought to provide the majority of the ligands to the Mn cluster in photosystem II. However, the precise ligands to the Mn in both the oxygen-evolving center and the Mn catalase have not yet

Figure 2 Proposed structures for the oxygen-evolving center (top) and Mn catalase active site (bottom), redrawn from Refs 14 and 18, respectively. The blue atoms are Mn, the red atoms are O and the yellow atoms are C. Some additional bonds to the Mn centers have been omitted for clarity.

been conclusively identified, so a definite conclusion as to whether a distant homology exists is not yet possible. Additional sequences from Mn catalases will be useful in this analysis. Was hydrogen peroxide present on the early Earth? The early atmosphere is thought to have been mildly reducing or neutral in overall redox balance1,2. Water photolysis by UV light can produce hydrogen peroxide, which then might be concentrated by rainfall in certain protected environments25. Thus, the presence of hydrogen peroxide on the early Earth is possible, although only a small amount might have accumulated due to its intrinsic reactivity. Therefore, it seems unlikely that hydrogen peroxide was ever the principal electron donor to the primitive photosystem because of the limited amounts available. A more likely candidate for an early electron donor is Fe2+, which was present in substantial quantities in the Archean oceans1. Purple photosynthetic bacteria that can oxidize ferrous iron to the ferric form have recently been described, although the enzyme complexes that catalyze this oxidation have not yet been identified26.

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TALKING POINT H 3C

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Figure 3 Chemical structures of (a) bacteriochlorophyll a and (b) chlorophyll a. Differences in the structures are shown in red. Chemical structures of (c) 3-acetyl-chlorophyll a and (d) chlorophyll d. R is the phytyl tail.

We propose that the Mn catalase developed to detoxify the hydrogen peroxide that was present in a local environment on the early Earth. This catalase was then recruited to extract electrons from the hydrogen peroxide, producing the first oxygen-evolving complex. Finally, the dimanganese center evolved to become the four-manganese center of the oxygen-evolving center so that it could extract electrons from water.

The development of a highly oxidizing reaction center While the scenario described above for the origin and evolution of the Mn center provides a plausible way in which the early reaction center might begin to produce oxygen from hydrogen peroxide, it does not address the issue of how the highly oxidizing species that is needed to split water developed. To understand this issue, it is important to appreciate that the essence of the primary electron transfer process in photosynthesis is an oxidation of the excited state of the reaction center primary donor pigment to generate a reduced acceptor molecule and an oxidized donor27. The effective redox potential of the excited state is a function of the ground state redox

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potential of the donor and the excitation energy. The ground state redox potential of the primary donor in purple bacterial reaction centers is approximately +0.5 V. This potential is determined by both the intrinsic redox behavior of the bacteriochlorophylls and the details of their environment in the reaction center protein. Recent work has shown that the redox potential of the primary donor bacteriochlorophyll a in reaction centers from the purple bacterium Rhodobacter sphaeroides can be raised dramatically by engineered mutations that increase the number of hydrogen bonds to the bacteriochlorophyll28. However, a necessary result of the increase in redox potential of the donor is that the potential of the excited state becomes less negative in parallel, because the redox span (i.e. the difference in redox potentials between the ground and excited states) is determined by the energy of the excitation photon, which is at 870 nm in Rb. sphaeroides (Fig. 1). If the excited state potential is increased too much, then the quantum yield of electron transfer falls, because the energy of the reduced acceptor now lies above the energy of the excited state of the donor. In fact, this has been shown to occur in engineered mutants in which

the potential of the donor has been greatly increased29. While changing the environment of the primary donor can substantially modulate the potential of the primary donor, that in itself is not enough to create a working reaction center with both a highly oxidizing potential of the donor and a sufficiently reducing potential of the excited state. The only way to do both these things is to increase the photon energy by shifting the absorption maximum from the near infrared into the visible region of the electromagnetic spectrum. This assumes that the redox properties of the electron acceptors are unchanged. The overall reaction center structure and especially the acceptor systems are very similar in photosystem II and the purple bacterial reaction centers. The chemical structures of chlorophyll a and bacteriochlorophyll a are shown in Fig. 3. There are two differences, which are shown in red. The first is the oxidation state of ring B, which is oxidized in chlorophyll a and reduced, converting the double bond into a single bond, in bacteriochlorophyll a. The second difference is the presence of the acetyl substituent at the 3 position in ring A in bacteriochlorophyll a instead of the vinyl found in chlorophyll a. The spectral differences between these two pigments are primarily due to the first of these two structural differences. How might bacteriochlorophyll a be converted into chlorophyll a? In the modern biosynthetic pathways of these pigments, chlorophyll is an intermediate on the way to bacteriochlorophyll30. The biosynthetic pathway of both these pigments includes a reduction of ring D, with the formation of a single bond instead of a double bond. In bacteriochlorophyll biosynthesis, an additional reduction of ring B takes place. This reaction is catalyzed by a set of three enzymes (coded by the bchlX, bchlY and bchlZ genes) that are clearly homologous to the enzymes (coded by the chlL, chlN and chlB genes) that reduce ring D in the more primitive oxygen-evolving organisms. Burke and coworkers31 proposed that the ancestral reductase was non-specific and reduced both rings B and D, producing bacteriochlorophyll without intermediate chlorophyll formation (however, see Ref. 32). A small change in substrate specificity of one or more of these primitive enzymes such that ring B is no longer reduced would produce a situation in which the chlorophyll-like pigment 3-acetyl-chlorophyll a is produced (Fig. 3c). This pigment is nearly identical to chlorophyll d, which has recently been reported as the

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Conclusions The proposed scenario is that the ability to oxidize hydrogen peroxide by an ancestral bacteriochlorophyll-containing reaction center similar to the purple bacterial complex was the first step in the progression that led to the oxygenevolving complex of photosystem II. The second step was the conversion from bacteriochlorophyll to chlorophyll, which raised the redox potential of the reaction center pigment sufficiently to oxidize the very weak electron donor water. Additional evolutionary steps that led to two linked photosystems were also needed to produce the present system found in all oxygenic photosynthetic organisms. These steps either could have followed the steps described above, or possibly could have preceded them.

Acknowledgements The authors thank Drs John Olson, James Allen and Wayne Frasch for helpful discussions. Supported by a grant from the Exobiology program of NASA. This is publication 342 from the Arizona State University Center for the Study of Early Events in Photosynthesis.

References 1 Schopf, J. W. and Klein, C., eds (1992) The Proterozoic Biosphere, Cambridge University Press 2 Kasting, J. F. (1993) Science 259, 920–926 3 Doolittle, R. F. et al. (1996) Science 271, 470–477 4 Castresana, J. and Saraste, M. (1995) Trends Biochem. Sci. 20, 443–448 5 Martin, W. F. (1996) BioEssays 18, 523–527

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principal pigment in a newly discovered oxygenic prokaryotic photosynthetic organism33. Chlorophyll d has an in vivo absorption maximum at only a slightly longer wavelength than chlorophyll a (716 nm versus 680 nm). This modest change in the biosynthetic enzyme specificity would produce a reaction center donor pigment that is structurally similar to bacteriochlorophyll but with a redox potential that is capable of oxidizing water. The acetyl group would fit the preexisting hydrogen-bonding interactions for the bacteriochlorophyll a. Finally, the ability to make the acetyl group at the 3 position was lost and the reaction center pigment became chlorophyll a. Information on the pathway and enzymes involved in the biosynthesis of chlorophyll d would be important in evaluating this proposal. Figure 4 shows the stages of development of the reaction center protein from purple bacterial complex to photosystem II. The intermediate stages involve the association of the Mn catalase and the intermediate pigment similar to chlorophyll d.

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Figure 4 Evolutionary stages of oxygen evolution capacity (OEC). Four stages are depicted, although additional intermediate stages undoubtedly also existed. For each stage, the upper diagram shows an energetic picture, and the lower diagram a schematic of the reaction center protein in the photosynthetic membrane. Stage 1: the energetics are the same as those found in contemporary purple bacterial reaction centers. The pigment is bacteriochlorophyll (BChl) a, and an external donor (D), possibly Fe2+, reduces the oxidized reaction center special pair pigment (P). Stage 2: the energetic and pigment composition are the same as stage 1, but the Mn catalase is present and can donate electrons to P, although probably with low efficiency. Stage 3: the pigment has changed to chlorophyll (Chl) d or a similar pigment, and the Mn catalase (Mn Cat) has become more closely associated with the reaction center. The higher excitation energy of the chlorophyll d pigment increases the redox potential of P. Stage 4: the pigment has converted to chlorophyll a and the Mn center has become fully incorporated into the reaction center protein. The known ground and excited state redox potentials for purple bacteria reaction centers were used for stages 1 and 2 and those of photosystem II for stage 4, calculated according to Ref. 27. The redox potentials of stage 3 were estimated using the in vivo excitation energy of chlorophyll d (Ref. 33), assuming that the redox potential of the excited state is unchanged from that of stage 2. 6 Ort, D. R. and Yocum, C. F., eds (1996) Oxygenic Photosynthesis: The Light Reactions, Kluwer 7 Olson, J. M. and Pierson, B. K. (1987) Int. Rev. Cytol. 108, 209–248 8 Blankenship, R. E. (1992) Photosynth. Res. 33, 91–111 9 Nitschke, W. and Rutherford, A. W. (1991) Trends Biochem. Sci. 16, 241–245 10 Mulkidjanian, A. Y. and Junge, W. (1997) Photosynth. Res. 51, 27–42 11 Whatley, J. M. (1993) Int. Rev. Cytol. 144, 259–299 12 Debus, R. J. (1992) Biochim. Biophys. Acta 1102, 269–352 13 Britt, R.D. (1996) in Oxygenic Photosynthesis: The Light Reactions (Ort, D. R. and Yocum, C. F., eds), pp. 137–164, Kluwer 14 Yachandra, V. K., Sauer, K. and Klein, M. P. (1996) Chem. Rev. 96, 2927–2950 15 Rögner, M., Boekema, E. J. and Barber, J. (1996) Trends Biochem. Sci. 44–49 16 Blankenship, R. E., Madigan, M. T. and Bauer, C. E., eds (1995) Anoxygenic Photosynthetic Bacteria, Kluwer 17 Olson, J. M. (1970) Science 168, 438–446 18 Penner-Hahn, J. E. (1992) in Manganese Redox Enzymes (Pecoraro, V., ed.), pp. 29–45, VCH 19 Dismukes, G. C. (1996) Chem. Rev. 96, 2909–2926 20 Rutherford, A. W. and Nitschke, W. (1996) in

21 22 23 24 25 26 27 28 29 30 31 32 33 34

Origin and Evolution of Biological Energy Conversion (Baltscheffsky, H., ed.), pp. 143–175, VCH Frasch, W. D. and Mei, R. (1987) Biochim. Biophys. Acta 891, 8–14 Bader, K. P. (1994) Biochim. Biophys. Acta 1188, 213–219 Samuilov, V. D. (1997) Biochemistry (Moscow) 62, 451–454 Igarashi, T., Kono, Y. and Tanaka, K. (1996) J. Biol. Chem. 271, 29521–29524 McKay, C. P. and Hartman, H. (1991) Origins Life Evol. Biosphere 21, 157–163 Widdle, F. et al. (1993) Nature 362, 834–836 Blankenship, R. E. and Prince, R. C. (1985) Trends Biochem. Sci. 10, 382–383 Lin, X. et al. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 10265–10269 Woodbury, N. W. et al. (1995) Chem. Phys. 197, 405–421 Porra, R. J. (1997) Photochem. Photobiol. 65, 492–516 Burke, D. H., Hearst, J. E. and Sidow, A. (1993) Proc. Natl. Acad. Sci. U. S. A. 90, 7134–7138 Lockhart, P. J. et al. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 1930–1934 Miyashita, H. et al. (1996) Nature 338, 402 George, P. (1965) in Oxidases and Related Redox Systems (Vol. 1) (King, T. E., Mason, H. S. and Morrison, M., eds), pp. 3–36, Wiley

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