576th MEETING, LONDON
901
PHOTOSYSTEM II Colloquium organized on behalf of the Bioenergetics Group by J. W. Bradbeer (London) and 0. T. G. Jones (Bristol) Oxygen Evolution: Some Historical Aspects ROBERT HILL Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1 Q W, U.K. Are we all safe now to be celebrating the 40th anniversary of the experimental separation of the production of O2 in light from the reduction of C 0 2 ? The answer in the affirmative was really given nearly 50 years ago by Kluyver and van Niel in The Netherlands. The production of O2 by reaction 11 could then represent the first chemical process which was shown in vitro to belong to photosynthesis. The production of molecular O2 depended on the reduction of a hydrogen acceptor so that water could be regarded as the photochemical reactant. This provided direct experimental suprort for the complete scheme of van Niel, which originated from his studies with the photosynthetic bacteria (van Niel, 1949). Haemoglobin and chlorophyll The dominant ideas for the interpretation of photosynthesis during the first quarter
of the present century were fully described by H. A. Spoehr (1926); his book gives a detailed account of the state of knowledge at that time. There were, it seems, two great influences: F. F. Blackman (Spoehr, 1926, page 95) by means of his analysis of the physiological process and Willstatter by his chemical analysis of the chlorophyll molecule (Spoehr, 1926, page 355). The green colour of the grass and the red colour of the blood had been shown earlier by Hoppe-Seyler and others to belong to closely related chemical compounds. Willstatter had discovered that chlorophyll was a compound of magnesium with a tetrapyrrolic residue. This seemed to give to the plant pigment a rank equal to the iron-containing respiratory pigment haemoglobin. In 1909 R. A. Peters had discovered that one atom of iron corresponded to one molecule of 0, when the haernoglobin was saturated with the gas. Oxyhaemoglobin was given the status of a definite chemical compound. This could be interpreted by the theory of Werner concerning the molecular compounds as referring to co-ordination numbers with complex salts of metals. Chemical theory of Willstatter and SroN Willstatter had been very much under the influence of Baeyer. The theory (Spoehr, 1926, page 282) was based partly on Baeyer’s formaldehyde theory together with the special properties postulated for the magnesium atom in chlorophyll. The photochemical process was given as the rearrangement of the atoms in bicarbonate when it was combined with the magnesium atom in the chlorophyll. This was represented firstly as the conversion into a formyl peroxide and secondly as a conversion into a formaldehyde peroxide compound of chlorophyll. The formaldehyde was to be liberated from the chlorophyll after the removal of the peroxide giving molecular 0 2Chlorophyll . in fact was found not to give a compound with formaldehyde. The decomposition of the peroxide then represented the enzymic dark reaction as referred to by Blackman. Although this theory was far from being verified in vitro it was so simple that it gave a readily acceptable picture of photosynthesis. At that time the presence of COz was essential for the production of O2 by a green plant in light.
Vol. 6
902
BIOCHEMICAL SOCIETY TRANSACTIONS
Chlorella and photosynthesis Otto Warburg initiated a revolution in the study of photosynthesis. He made manometric determinations of the gas exchange with suspensions of the green alga Chlorella (Krebs, 1972). The action of inhibitors was determined under different conditions of illumination. Cyanide, the ‘metal poison’, was shown to inhibit the dark reaction (in Blackman’s sense). The narcotic poisons represented by the urethanes inhibited the photochemical process. The quantum theory developed by Max Planck in 1900 was soon to be applied to photochemistry. Einstein in 1912 had stated the law of photochemical equivalence. The application of this law by Warburg to the interpretation of photosynthesis was a natural consequence of his being at the focus of events in Berlin. Biochemical theory of Kieta Shibata In 1931, lectureson photosynthesis by KietaShibata had been published in Japan.They have recently been republished posthumously with a translation in English (Takamiya, 1975). They give a clear and concise account of the knowledge at that time. The author gave his own theory of carbon assimilation; bicarbonate, as in the theory of Willstatter, was bound to the magnesium by one of the electrovalencies; the magnesium, according to Shibata, also took up four water molecules; instead of the photochemical rearrangement of the bicarbonate the water molecules were broken down to H and OH; the four hydrogen atoms reduced the bicarbonate and the four hydroxyl groups were to give H202. The inhibition of the dark reaction by cyanide indicated an essential participation of catalase, which became a lively issue, especially in Japan. Biochemical studies with haemoglobin My own direct contact with photosynthesis was both accidental and spontaneous; it resulted from a study of the blood pigment over the period 1923-1938 and was conditioned by my liking for colours and for Werner’s work on co-ordination compounds. This preoccupation with haemoglobin led to an association with D. Keilin after his first paper on the function of cytochrome in respiration was published. I was deeply involved in his work on the separation of cytochrome c from the complete system in baker’s yeast. The functional analysis of the cytochrome system by Keilin (1966, page 164) had settled a controversy concerning respiration. The components of cytochrome acted as a link between O2 and the transfer of hydrogen from metabolites. The reduction of the cytochrome components was inhibited by narcotics and the oxidation by the metal poisons. Haemoglobin for measuring pressures of O2 Between 1933 and 1935 I was in contact with Joseph Barcroft in connection with variation in oxygen affinity shown by different haemoglobins. This was concerned with my use of a method for determining an oxygen dissociation-curve with a small quantity of haemoglobin. The discovery by Keilin (1966, page 176) of the oxidation of cytochrome c with the so-called indophenol oxidase showed how the cytochrome system could function at very low pressures of 02,the high affinity of myoglobin at low O2pressures and the hypobolic dissociation curve showed adaptation to its physiological function in transferring O2from the circulation to a tissue. Hence after 10 years acquaintance with Spoehr’s (1926) book one was tempted to try the use of myoglobin for detecting liberation of O2 by isolated chloroplasts. Measurable oxygen production from isolated chloroplasts I had previously examined a number of plants with the object of obtaining suspensions of chloroplasts, but none of these showed any signs of giving a measurable O2 output with the myoglobin method. The addition of a soluble factor from the leaf to represent the enzymic dark factor of Blackman was tried. This gave a measurable effect and formed the starting point. The finding that the presence or relative absence of 1978
576th MEETING, LONDON
903
carbonate made no differencewas originally most disappointing. The active agent seemed to be iron compounds of organic acids and led to the use of ferric potassium oxalate, which was compatible with haemoglobin. This provided convincing evidence that the isolated chloroplasts would catalyse the photochemical reduction of the ferric iron according to an equation now to be given by:
+
+
+ +
4K3Fe(C204)3 2H20 = 4K2Fe(C204)2 HzC204 O2 304kJ The energy per einstein for the light absorbed by the chlorophyll would be about 170kJ (40kcal). This on a one quantum per electron basis represented an efficiencyof about 45 % if the first quantum yield measurements of Warburg were accepted. Separation of O2productionfrom COz reduction The most important point was to see if inhibitors of the enzyme catalase would affect the O,-producing reaction of the chloroplasts. It was fortunate that the concentrations of the so-called metal poisons, which would completely inhibit catalase, gave no reaction with either the haemoglobin or with the ferric-ferro oxalate. The photochemical liberation of 0,was found not to depend upon catalase acting upon H202. It remained to show whether the chloroplast reaction was in fact a part of the photosynthetic process. This problem owed its solution to results obtained by Warburg with Chlorella. R. Scarisbrick and I found that the chloroplast reaction was inhibited by the urethanes at the concentrations which had been shown to inhibit photosynthesis in Chlorella. The production of oxygen then must correspond to the light reaction in the sense used by Blackman. In 1939 Ruben, Hassid and Kamen (Kamen, 1949) published the first account of following the assimilation process with "CO,. This seemed, happily, to divide the study of photosynthesis into the two separate kinds of endeavour as indicated by Blackman for the photochemical and the enzymic processes. The studies with isolated chloroplasts were continued by C. S. French and colleagues (Holt & French, 1949). French succeeded H. A. Spoehr as Director of the Carnegie Institution of Washington, Standord, and formed an international centre for biochemical and biophysical studies of photosynthesis. Redox system in chloroplasts When it came to the analysis of the 0,-producing system of the green plant the analogy of a cytochrome system driven in reverse immediately suggested itself. This at once led to the search for cytochrome components in chloroplasts, and component f was characterized. I was very fortunate to have the association with R. Scarisbrick in connection with this. We had a really exciting interval of about 3 years. Then came the Second World War and we both moved to other occupations. The studies of cytochrome components were not published until 1950. Yet in those few years it seems that the foundations for the biochemical study of photosynthesis had been securely laid. The most outstanding fact was the extreme lability of the 02-producing system. The 'red drop' interpreted by Emerson and Rabinovitch (Takamiya, 1975, page 140) After 1939 the question of the quantum requirement, 1/4, had become a controversial issue. Many careful determinations in different departments had given a requirement of more than twice that originally found by Warburg. Robert Emerson devoted years of endeavour, which maintained the evidence for the higher requirement. Furthermore Emerson and Lewis determined l / d as a function of the wavelength of the light. The comparison of the action spectrum with the absorption spectrum gave an unexplained fall in the quantum efficiency, 4, at the red end of the spectrum. Emerson and Chalmers found that this fall in 4 could be eliminated by simultaneous illumination with a shorter wavelength. It was concluded that two photochemical systems were required; this gave a new interpretation for results obtained with different algae in other laboratories.
Vol. 6
904
BIOCHEMICAL SOCIETY TRANSACTIONS
Origin of the ‘Z scheme’
Severo Ochoa (1951) had suggested that the reduction of CO, could depend on the photochemical reduction of NADP+. This was proved by Vishniac and Ochoa with a chloroplast preparation supplied with a hydrogen donor at a less oxidizing potential than water. The potential span required for photosynthesis would be near to that of the oxyhydrogen reaction, from +0.32V to -0.41 V at pH7. The characteristic potential of cytochromefwas determined by H. E. Davenport as +0.35V at pH7. D. S. Bendall had found that a b-type cytochrome present in chloroplasts had a characteristic potential of +O at pH7. It seemed that the postulation of two forward reactions joined by a back reaction would rationalize the distribution of potentials. This hypothetical scheme, later referred to as the Z scheme, was developed with F. Bendall (Takamiya, 1975, page 55). The scheme survived because the potentials of other components of the chloroplast were found to be near to one or other of the two cytochrome components (Hill, 1965). Hill, R. (1965) Essays Biochern. 1, 121-151
Holt, A. S. & French, C. S. (1 949) in Photosynthesis inPlanrs (Franck,J. & Loomis, W. E., eds.), pp. 277-285, Iowa State College Press, Iowa Kamen, M. D. (1949) in PhotosynthesisinPlants(Franck, J. & Loomis, W. E.,eds.), pp. 365-380, Iowa State College Press, Iowa Keilin, D. (1966) in The History of Cell Respiration and Cytochrome, Cambridge University Press, Cambridge Krebs, H. A. (1972) Biogr. Mem. Fellows R . SOC.18,629-652 Ochoa, S . (1951) Exp. Biol. 5,29-51 Spoehr, H. A. (1926) Photosynthesis, Chemical Catalog Company, New York Takamiya, A. (ed.) (1975) Translation of Carbon and Nitrogen Assimilation by Kieta Shibata (1931), Japan Science Press, Tokyo van Niel, C. B. (1949) inphotosynthesis inPlunfs(Franck,J. & Loomis, W. E., eds.), pp. 437-495, Iowa State College Press, Iowa
Variations in Photosynthetic Electron-Transport Pathways in Chlorella vulgaris JOHN SINCLAIR and AKINORI SARA1 Department of Biology, Carlton University, Ottawa, Ontario, Canada Photosynthesis in green plants and algae is known to involve O2 as both a product and a reactant. It is produced when water molecules are consumed in a reaction that feeds electrons into the photosynthetic electron-transport pathway. 0,is consumed during photorespiration (see review by Jackson & Volk, 1970), a light driven process that results in the release of CO,. This process appears to be intimately connected with the functioning of the enzyme ribulose 1,5-bisphosphate carboxylase-oxygenase (EC 4.1.1.39). O2 can also cause an inhibition of C 0 2fixation (Warburg effect), although the mechanism of this effect appears to be another aspect of photorespiration. O2can be reduced to H 2 0 2by Photosystem I in the Mehler reaction, which was first demonstrated by Mehler (1951) with isolated chloroplasts. The demonstration by Arnon et a/. (1967) that O2 could be reduced by ferredoxin in the Mehler reaction gave support to the idea that this reaction operated in uiuo. Evidence for the Mehler reaction in intact algal cells came from the work of Patterson & Myers (1973) with Anacysfisnidulans. They showed that H 2 0 2 was produced in a light-driven process, which was inhibited by 3-(3,4dichloropheny1)-1,I-dimethylurea. This process operated more rapidly at high O2 concentrations and low C 0 2 concentrations. Evidence that O2 could accept electrons from a site between the two photosystems was provided by Diner & Mauzerall (1973). The latter studied the evolution of 0,from Chlorella vulgaris and Phormidium luridum in the presence of different concentrations of 0,.They found that at low 0, 1978
901
PHOTOSYSTEM II Colloquium organized on behalf of the Bioenergetics Group by J. W. Bradbeer (London) and 0. T. G. Jones (Bristol) Oxygen Evolution: Some Historical Aspects ROBERT HILL Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1 Q W, U.K. Are we all safe now to be celebrating the 40th anniversary of the experimental separation of the production of O2 in light from the reduction of C 0 2 ? The answer in the affirmative was really given nearly 50 years ago by Kluyver and van Niel in The Netherlands. The production of O2 by reaction 11 could then represent the first chemical process which was shown in vitro to belong to photosynthesis. The production of molecular O2 depended on the reduction of a hydrogen acceptor so that water could be regarded as the photochemical reactant. This provided direct experimental suprort for the complete scheme of van Niel, which originated from his studies with the photosynthetic bacteria (van Niel, 1949). Haemoglobin and chlorophyll The dominant ideas for the interpretation of photosynthesis during the first quarter
of the present century were fully described by H. A. Spoehr (1926); his book gives a detailed account of the state of knowledge at that time. There were, it seems, two great influences: F. F. Blackman (Spoehr, 1926, page 95) by means of his analysis of the physiological process and Willstatter by his chemical analysis of the chlorophyll molecule (Spoehr, 1926, page 355). The green colour of the grass and the red colour of the blood had been shown earlier by Hoppe-Seyler and others to belong to closely related chemical compounds. Willstatter had discovered that chlorophyll was a compound of magnesium with a tetrapyrrolic residue. This seemed to give to the plant pigment a rank equal to the iron-containing respiratory pigment haemoglobin. In 1909 R. A. Peters had discovered that one atom of iron corresponded to one molecule of 0, when the haernoglobin was saturated with the gas. Oxyhaemoglobin was given the status of a definite chemical compound. This could be interpreted by the theory of Werner concerning the molecular compounds as referring to co-ordination numbers with complex salts of metals. Chemical theory of Willstatter and SroN Willstatter had been very much under the influence of Baeyer. The theory (Spoehr, 1926, page 282) was based partly on Baeyer’s formaldehyde theory together with the special properties postulated for the magnesium atom in chlorophyll. The photochemical process was given as the rearrangement of the atoms in bicarbonate when it was combined with the magnesium atom in the chlorophyll. This was represented firstly as the conversion into a formyl peroxide and secondly as a conversion into a formaldehyde peroxide compound of chlorophyll. The formaldehyde was to be liberated from the chlorophyll after the removal of the peroxide giving molecular 0 2Chlorophyll . in fact was found not to give a compound with formaldehyde. The decomposition of the peroxide then represented the enzymic dark reaction as referred to by Blackman. Although this theory was far from being verified in vitro it was so simple that it gave a readily acceptable picture of photosynthesis. At that time the presence of COz was essential for the production of O2 by a green plant in light.
Vol. 6
902
BIOCHEMICAL SOCIETY TRANSACTIONS
Chlorella and photosynthesis Otto Warburg initiated a revolution in the study of photosynthesis. He made manometric determinations of the gas exchange with suspensions of the green alga Chlorella (Krebs, 1972). The action of inhibitors was determined under different conditions of illumination. Cyanide, the ‘metal poison’, was shown to inhibit the dark reaction (in Blackman’s sense). The narcotic poisons represented by the urethanes inhibited the photochemical process. The quantum theory developed by Max Planck in 1900 was soon to be applied to photochemistry. Einstein in 1912 had stated the law of photochemical equivalence. The application of this law by Warburg to the interpretation of photosynthesis was a natural consequence of his being at the focus of events in Berlin. Biochemical theory of Kieta Shibata In 1931, lectureson photosynthesis by KietaShibata had been published in Japan.They have recently been republished posthumously with a translation in English (Takamiya, 1975). They give a clear and concise account of the knowledge at that time. The author gave his own theory of carbon assimilation; bicarbonate, as in the theory of Willstatter, was bound to the magnesium by one of the electrovalencies; the magnesium, according to Shibata, also took up four water molecules; instead of the photochemical rearrangement of the bicarbonate the water molecules were broken down to H and OH; the four hydrogen atoms reduced the bicarbonate and the four hydroxyl groups were to give H202. The inhibition of the dark reaction by cyanide indicated an essential participation of catalase, which became a lively issue, especially in Japan. Biochemical studies with haemoglobin My own direct contact with photosynthesis was both accidental and spontaneous; it resulted from a study of the blood pigment over the period 1923-1938 and was conditioned by my liking for colours and for Werner’s work on co-ordination compounds. This preoccupation with haemoglobin led to an association with D. Keilin after his first paper on the function of cytochrome in respiration was published. I was deeply involved in his work on the separation of cytochrome c from the complete system in baker’s yeast. The functional analysis of the cytochrome system by Keilin (1966, page 164) had settled a controversy concerning respiration. The components of cytochrome acted as a link between O2 and the transfer of hydrogen from metabolites. The reduction of the cytochrome components was inhibited by narcotics and the oxidation by the metal poisons. Haemoglobin for measuring pressures of O2 Between 1933 and 1935 I was in contact with Joseph Barcroft in connection with variation in oxygen affinity shown by different haemoglobins. This was concerned with my use of a method for determining an oxygen dissociation-curve with a small quantity of haemoglobin. The discovery by Keilin (1966, page 176) of the oxidation of cytochrome c with the so-called indophenol oxidase showed how the cytochrome system could function at very low pressures of 02,the high affinity of myoglobin at low O2pressures and the hypobolic dissociation curve showed adaptation to its physiological function in transferring O2from the circulation to a tissue. Hence after 10 years acquaintance with Spoehr’s (1926) book one was tempted to try the use of myoglobin for detecting liberation of O2 by isolated chloroplasts. Measurable oxygen production from isolated chloroplasts I had previously examined a number of plants with the object of obtaining suspensions of chloroplasts, but none of these showed any signs of giving a measurable O2 output with the myoglobin method. The addition of a soluble factor from the leaf to represent the enzymic dark factor of Blackman was tried. This gave a measurable effect and formed the starting point. The finding that the presence or relative absence of 1978
576th MEETING, LONDON
903
carbonate made no differencewas originally most disappointing. The active agent seemed to be iron compounds of organic acids and led to the use of ferric potassium oxalate, which was compatible with haemoglobin. This provided convincing evidence that the isolated chloroplasts would catalyse the photochemical reduction of the ferric iron according to an equation now to be given by:
+
+
+ +
4K3Fe(C204)3 2H20 = 4K2Fe(C204)2 HzC204 O2 304kJ The energy per einstein for the light absorbed by the chlorophyll would be about 170kJ (40kcal). This on a one quantum per electron basis represented an efficiencyof about 45 % if the first quantum yield measurements of Warburg were accepted. Separation of O2productionfrom COz reduction The most important point was to see if inhibitors of the enzyme catalase would affect the O,-producing reaction of the chloroplasts. It was fortunate that the concentrations of the so-called metal poisons, which would completely inhibit catalase, gave no reaction with either the haemoglobin or with the ferric-ferro oxalate. The photochemical liberation of 0,was found not to depend upon catalase acting upon H202. It remained to show whether the chloroplast reaction was in fact a part of the photosynthetic process. This problem owed its solution to results obtained by Warburg with Chlorella. R. Scarisbrick and I found that the chloroplast reaction was inhibited by the urethanes at the concentrations which had been shown to inhibit photosynthesis in Chlorella. The production of oxygen then must correspond to the light reaction in the sense used by Blackman. In 1939 Ruben, Hassid and Kamen (Kamen, 1949) published the first account of following the assimilation process with "CO,. This seemed, happily, to divide the study of photosynthesis into the two separate kinds of endeavour as indicated by Blackman for the photochemical and the enzymic processes. The studies with isolated chloroplasts were continued by C. S. French and colleagues (Holt & French, 1949). French succeeded H. A. Spoehr as Director of the Carnegie Institution of Washington, Standord, and formed an international centre for biochemical and biophysical studies of photosynthesis. Redox system in chloroplasts When it came to the analysis of the 0,-producing system of the green plant the analogy of a cytochrome system driven in reverse immediately suggested itself. This at once led to the search for cytochrome components in chloroplasts, and component f was characterized. I was very fortunate to have the association with R. Scarisbrick in connection with this. We had a really exciting interval of about 3 years. Then came the Second World War and we both moved to other occupations. The studies of cytochrome components were not published until 1950. Yet in those few years it seems that the foundations for the biochemical study of photosynthesis had been securely laid. The most outstanding fact was the extreme lability of the 02-producing system. The 'red drop' interpreted by Emerson and Rabinovitch (Takamiya, 1975, page 140) After 1939 the question of the quantum requirement, 1/4, had become a controversial issue. Many careful determinations in different departments had given a requirement of more than twice that originally found by Warburg. Robert Emerson devoted years of endeavour, which maintained the evidence for the higher requirement. Furthermore Emerson and Lewis determined l / d as a function of the wavelength of the light. The comparison of the action spectrum with the absorption spectrum gave an unexplained fall in the quantum efficiency, 4, at the red end of the spectrum. Emerson and Chalmers found that this fall in 4 could be eliminated by simultaneous illumination with a shorter wavelength. It was concluded that two photochemical systems were required; this gave a new interpretation for results obtained with different algae in other laboratories.
Vol. 6
904
BIOCHEMICAL SOCIETY TRANSACTIONS
Origin of the ‘Z scheme’
Severo Ochoa (1951) had suggested that the reduction of CO, could depend on the photochemical reduction of NADP+. This was proved by Vishniac and Ochoa with a chloroplast preparation supplied with a hydrogen donor at a less oxidizing potential than water. The potential span required for photosynthesis would be near to that of the oxyhydrogen reaction, from +0.32V to -0.41 V at pH7. The characteristic potential of cytochromefwas determined by H. E. Davenport as +0.35V at pH7. D. S. Bendall had found that a b-type cytochrome present in chloroplasts had a characteristic potential of +O at pH7. It seemed that the postulation of two forward reactions joined by a back reaction would rationalize the distribution of potentials. This hypothetical scheme, later referred to as the Z scheme, was developed with F. Bendall (Takamiya, 1975, page 55). The scheme survived because the potentials of other components of the chloroplast were found to be near to one or other of the two cytochrome components (Hill, 1965). Hill, R. (1965) Essays Biochern. 1, 121-151
Holt, A. S. & French, C. S. (1 949) in Photosynthesis inPlanrs (Franck,J. & Loomis, W. E., eds.), pp. 277-285, Iowa State College Press, Iowa Kamen, M. D. (1949) in PhotosynthesisinPlants(Franck, J. & Loomis, W. E.,eds.), pp. 365-380, Iowa State College Press, Iowa Keilin, D. (1966) in The History of Cell Respiration and Cytochrome, Cambridge University Press, Cambridge Krebs, H. A. (1972) Biogr. Mem. Fellows R . SOC.18,629-652 Ochoa, S . (1951) Exp. Biol. 5,29-51 Spoehr, H. A. (1926) Photosynthesis, Chemical Catalog Company, New York Takamiya, A. (ed.) (1975) Translation of Carbon and Nitrogen Assimilation by Kieta Shibata (1931), Japan Science Press, Tokyo van Niel, C. B. (1949) inphotosynthesis inPlunfs(Franck,J. & Loomis, W. E., eds.), pp. 437-495, Iowa State College Press, Iowa
Variations in Photosynthetic Electron-Transport Pathways in Chlorella vulgaris JOHN SINCLAIR and AKINORI SARA1 Department of Biology, Carlton University, Ottawa, Ontario, Canada Photosynthesis in green plants and algae is known to involve O2 as both a product and a reactant. It is produced when water molecules are consumed in a reaction that feeds electrons into the photosynthetic electron-transport pathway. 0,is consumed during photorespiration (see review by Jackson & Volk, 1970), a light driven process that results in the release of CO,. This process appears to be intimately connected with the functioning of the enzyme ribulose 1,5-bisphosphate carboxylase-oxygenase (EC 4.1.1.39). O2 can also cause an inhibition of C 0 2fixation (Warburg effect), although the mechanism of this effect appears to be another aspect of photorespiration. O2can be reduced to H 2 0 2by Photosystem I in the Mehler reaction, which was first demonstrated by Mehler (1951) with isolated chloroplasts. The demonstration by Arnon et a/. (1967) that O2 could be reduced by ferredoxin in the Mehler reaction gave support to the idea that this reaction operated in uiuo. Evidence for the Mehler reaction in intact algal cells came from the work of Patterson & Myers (1973) with Anacysfisnidulans. They showed that H 2 0 2 was produced in a light-driven process, which was inhibited by 3-(3,4dichloropheny1)-1,I-dimethylurea. This process operated more rapidly at high O2 concentrations and low C 0 2 concentrations. Evidence that O2 could accept electrons from a site between the two photosystems was provided by Diner & Mauzerall (1973). The latter studied the evolution of 0,from Chlorella vulgaris and Phormidium luridum in the presence of different concentrations of 0,.They found that at low 0, 1978
