Light-dependent hydrogen evolution by Scenedesmus

Using manometric and enzymic techniques, H2 and CO2 evolution in darkness and light has been studied in the green alga Chianydomonas reinhardtii F-60. F-60 is a mutant strain characterized by an incomplete photosynthetic carbon reduction cycle but an intact electron transport chain.In the dark, starch was broken down, and H2 and CO2 was released. The uncoupler, carbonyl cyanide m-fluorophenylhydrazone with an optimum concentration of 5 to 10 micromolar, increased the rate of CO2 release and starch breakdown but depressed H2 formation. It m-fluorophenylhydrazone, stimulated H2 photoproduction by removing ATP which limited the sequence of reactions. The contribution of photosystem II to the photoproduction of H2, as judged from the effect of 10 micromolar 3-(3,4-dichlorophenyl)-1,1-dimethylurea, was at least 80%.CO2 photoevolution increased Unearly with time, but H2 photoevolution occurred in two phases: a rapid initial phase folowed by a second slower phase. The rate of H2 release increased hyperbolically with light intensity, but the rate of CO2 production tended to level off and decrease with increasing light intensity, up to 145 watts per square meter. It was proposed that a changing CO2 and H2 ratio is the result of interaction between the carbon and hydrogen metabolism and the photosynthetic electron transport chain.Studies with chlorophyllous algae have established that H2 evolution in the light is the result of electron transport through the photosystems associated with an adaptable hydrogenase. These algae also produce H2 in the dark but at a lower rate. Two mechanisms have been proposed to account for H2 release in the light, while the pathway for the dark release of H2 has received 'Supported by Department of Energy (10-EY-76-5-02-3231) and National Science Foundation (PCM 79-22612 light (3, 14). It is known that gas evolution is stimulated by added glucose (5). Data obtained with position-labeled glucose led Kaltwasser et al. (13) to propose that classical glycolysis in Scenedesmus obliquus is the pathway involved. From a stoichiometric analysis of the products-which included ethanol, glycerol, and acetate, in addition to CO2 and H2-Klein and Betz (15) also proposed glycolysis for the heterofermentative breakdown of the reserve substance, starch, in Chlamydomonas reinhardii. This suggestion was supported by a similar fermentation pattern in Chlorella vulgaris (22). On the basis of a depression of H2 evolution in the dark by uncouplers of phosphorylation, Gaffron and Rubin (5) suggested that the fermentation yielded not only CO2 but also ATP. ATP would be required to raise the redox potential of the electrons from the reductant (NADH) to a higher one necessary for H2 production. In contrast to inhibiting H2 evolution in the dark, the uncoupler elevated the photorelease of H2 and CO2. Acetate has been reported to stimulate H2 evolution insensitive to DCMU, and Healy (10) suggested an anaerobically functioning citric acid cycle in Chlamydomonas moewusii to explain this observation. In his formu...

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confidence: 99%

Section: Introductionmentioning

confidence: 99%

H₂ and CO₂ Evolution by Anaerobically Adapted Chlamydomonas reinhardtii F-60

Bamberger¹,

King²,

Erbes³

et al. 1982

“…Another aspect of interest here is the anaerobic stimulatory effect of acetate on the release of CO2 and H2 in the light by C. reinhardii (3) and by other green algae ( 18,21 acetate requires ATP, Bamberger et al (3) suggested that acetate increased gas production by consuming ATP which regulated the unspecified sequence of reactions giving rise to CO2 and H2. Healey (18) modifying a mechanism put forward by Jones and Myers (20) to explain the Kok effect in blue-green algae, proposed a flow of electrons from acetate via the citric acid cycle into PSI resulting in the photoevolution of H2 from reduced Fd.…”

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confidence: 99%

Fermentative Metabolism of Chlamydomonas reinhardii

Gibbs¹,

Gfeller²,

Chen³

1986

The anaerobic photodissimilation of acetate by Chlamydomonas reinhardii F-60 adapted to a hydrogen metabolism was studied utilizing manometric and isotopic techniques. The rate of photoanaerobic (N2) acetate uptake was approximately 20 moles per milligram chlorophyll per hour or one-half that of the photoaerobic (air) rate. Under N2, cells produced 1.7 moles H2 and 0.8 mole CO2 per mole of acetate consumed. Gas production and acetate uptake were inhibited by monofluoroacetic acid (MFA), 3-(3',4'-dichlorophenyl)-1,1-dimethylurea (DCMU) and by H2. Acetate uptake was inhibited about 50% by 5% H2 (95% N2 (3) suggested that acetate increased gas production by consuming ATP which regulated the unspecified sequence of reactions giving rise to CO2 and H2. Healey (18) modifying a mechanism put forward by Jones and Myers (20) to explain the Kok effect in blue-green algae, proposed a flow of electrons from acetate via the citric acid cycle into PSI resulting in the photoevolution of H2 from reduced Fd. The operation of an anaerobic and light-dependent citric acid cycle which affects the stoichiometric conversion of acetate to CO2 and H2 had been documented in the photosynthetic purple bacteria ( 13).The present communication summarizes the results of a detailed investigation of the anaerobic photometabolism of acetate by C. reinhardii F-60, with reference to stoichiometry of gas (CO2 and H2) production, incorporation into cellular components, and sensitivity of the process to a variety of inhibitors. The stoichiometric relationships observed, together with the isotopic distribution following assimilation of ["4C]acetate, constitute strong evidence for the conclusion that anaerobic carbon oxidation occurs in part through the reactions of the glyoxylate and citric acid cycles. MATERIALS AND METHODSAlgal Growth Conditions. Chlamydomonas reinhardii (Dangeard) F-60, a mutant strain with an incomplete photosynthetic carbon reduction cycle but with an intact photosynthetic electron transport chain, was obtained from R. K. Togasaki, Indiana University. Cells were grown in batch cultures on an acetatesupplemented medium (14)

“…Classical glycolysis followed by subsequent metabolism of the pyruvate to the various end-products has been proposed to account for the anaerobic catabolism of starch or glucose in the green algae (17,19,27). Except for H2 evolution, the effect of light on fermentative carbon flow has received little attention.…”

Section: Introductionmentioning

confidence: 99%

“…In all conditions in the light, ethanol is a minor product. Formate production is least affected by light.The stoichiometry in the dark indicates that starch is degraded via the glycolytic pathway, and pyruvate is broken down into acetyl- (3,19,20,28).Classical glycolysis followed by subsequent metabolism of the pyruvate to the various end-products has been proposed to account for the anaerobic catabolism of starch or glucose in the green algae (17,19,27). Except for H2 evolution, the effect of light on fermentative carbon flow has received little attention.…”

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confidence: 99%

Fermentative Metabolism of Chlamydomonas reinhardtii

Gfeller¹,

Gibbs²

1984

245

128

The anaerobic starch breakdown into end-products in the green alg Chiamydomonas reinhardtii F-60 has been investigted in the dark and in the light. The effects of 343,4-dichlorophenyl)-1,1-dimethylurea (DCMU) and carbonyl cyanide-p-trifluoromethoxyphenyl hydrazone (FCCP) on the fermentation in the light have also been investigted.Anaerobic starch breakdown rnte (13.1 ± 3.5 micromoles C per milligram chlorophyll per hour) is increased 2-fold by FCCP in the dark. Light (100 watts per square meter) decreases up to 4-fold the dark rate, an inhibition reversed by FCCP. Stimulation of starch breakdown by the proton ionophore FCCP points to a pH-controlled rate-limiting step in the dark, while inhibition by light, and its reversal by FCCP, indicates a control by energy charge in the light.In the dark, formate, acetate, and ethanol are formed in the ratios of 2.07:1.07:0.91, and account for roughly 100% of the C from the starch.H2 production is OA3 mole per mole glucose in the starch. Glycerol >-lactate, and CO2 have been detected in minor amounts.In the light, with DCMU and FCCP present, acetate is produced in a 1:1 ratio to formate, and H2 evolution is 2.13 moles per mole glucose. When FCCP only is present, acetate production is lower, and CO2 and H2 evolution is 1.60 and 4.73 moles per mole glucose, respectively.When DCMU alone is present, CO2 and H2 photoevolution is higher than in the dark. Without DCMU, CO2 and H2 evolution is about 100% higher than in its presence. In both conditions, acetate is not formed. In all conditions in the light, ethanol is a minor product. Formate production is least affected by light.The stoichiometry in the dark indicates that starch is degraded via the glycolytic pathway, and pyruvate is broken down into acetyl- (3,19,20,28).Classical glycolysis followed by subsequent metabolism of the pyruvate to the various end-products has been proposed to account for the anaerobic catabolism of starch or glucose in the green algae (17,19,27). Except for H2 evolution, the effect of light on fermentative carbon flow has received little attention. This is principally due to the problems involved with photosynthetic fixation of the CO2 that might be evolved fermentatively. The first to attempt to resolve this question were Klein and Betz (20) who reported that light had no effect on the rate of starch breakdown or the pattern of fermentation in Chlamydomonas moewusii. But they used extremely low levels of light (160 lux (2) made use of C. reinhardtii F-60, a mutant characterized by an incomplete photosynthetic carbon reduction pathway but an intact photosynthetic electron transport chain, to monitor 'true' CO2 and H2 evolution. To account for their results, they proposed an involvement between anaerobic carbohydrate metabolism and the photosynthetic electron transport chain, implying that carbohydrate degradation is entirely or partially localized in the chloroplast. The purpose of our study was to establish for the first time a complete fermentative balance in C. reinhardtii F-60 between...