Reductive carboxylate cycle

Bacterial ferredoxins function primarily as electron carriers in ferredoxin-mediated oxidation reduction reactions. Some examples are reduction of NAD, NADP, FMN, FAD, sulfite and protons in anaerobic bacteria, CO -fixation cycles in photosynthetic bacteria, nitrogen fixation in anaerobic nitrogen fixing bacteria, and reductive carboxylation of substrates in fermentative bacteria. The roles of bacterial ferredoxins in these reactions have been summarized by Orme-Johnson (2), Buchanan and Arnon (3), and Mortenson and Nakos (31). 

Figure 17-14 (A) The reductive carboxylation system used in reductive pentose phosphate pathway (Calvin-Benson cycle). The essential reactions of this system are enclosed within the dashed box. Typical subsequent reactions follow. The phosphatase action completes the phosphorylation-dephosphorylation cycle. (B) The reductive pentose phosphate cycle arranged to show the combining of three C02 molecules to form one molecule of triose phosphate. Abbreviations are RCS, reductive carboxylation system (from above) A, aldolase, Pase, specific phosphatase and TK, transketolase.

The question is therefore, what are the principal requirements of an autotrophic carbon-fixation mechanism An organic molecule serves as a C02 acceptor molecule, which becomes carboxylated by a carboxylase enzyme. This C02 acceptor molecule needs to be regenerated in a reductive autocatalytic cycle. The product that can be drained off from such a metabolic cycle should be a central cellular metabolite, from which all cellular building blocks for polymers can be derived examples of such central metabolites are acetyl-CoA, pyruvate, oxaloacetate, 2-oxoghitarate, phosphoe-nolpyruvate, and 3-phosphoglycerate. Importantly, the intermediates should not be toxic to the cell. The irreversible steps of the pathway are driven by ATP hydrolysis, while the reduction steps are driven by low-potential reduced coenzymes. 

Three modifications of the conventional oxidative citric acid cycle are needed, which substitute irreversible enzyme steps. Succinate dehydrogenase is replaced by fumarate reductase, 2-oxoglutarate dehydrogenase by ferredoxin-dependent 2-oxoglutarate oxidoreductase (2-oxoglutarate synthase), and citrate synthase by ATP-citrate lyase [3, 16] it should be noted that the carboxylases of the cycle catalyze the reductive carboxylation reactions. There are variants of the ATP-driven cleavage of citrate as well as of isocitrate formation [7]. The reductive citric acid... 

The dicarboxylate/4-hydroxybutyrate cycle starts from acetyl-CoA, which is reductively carboxylated to pyruvate. Pyruvate is converted to PEP and then car-boxylated to oxaloacetate. The latter is reduced to succinyl-CoA by the reactions of an incomplete reductive citric acid cycle. Succinyl-CoA is reduced to 4-hydroxybu-tyrate, the subsequent conversion of which into two acetyl-CoA molecules proceeds in the same way as in the 3-hydroxypropionate/4-hydroxybutyrate cycle. The cycle can be divided into part 1 transforming acetyl-CoA, one C02 and one bicarbonate to succinyl-CoA via pyruvate, PEP, and oxaloacetate, and part 2 converting succinyl-CoA via 4-hydroxybutyrate into two molecules of acetyl-CoA. This cycle was shown to function in Igrticoccus hospitalis, an anaerobic autotrophic hyperther-mophilic Archaeum (Desulfurococcales) [40]. Moreover, this pathway functions in Thermoproteus neutrophilus (Thermoproteales), where the reductive citric acid cycle was earlier assumed to operate, but was later disproved (W.H. Ramos-Vera et al., unpublished results). 

Evans, Buchanan, and Arnon (41a) have recently found that the ferredoxin-dependent pyruvate and a-ketoglutarate synthesizing reactions function in a new carbon cycle for the photosynthetic fixation of C02. The new cycle was named the reductive carboxylic acid cycle, and apart from pyruvate and a-ketoglutarate synthases, it includes certain of the enzymes associated with Krebs citric acid cycle, operating in the synthetic direction. Photoreduced ferredoxin and ATP, formed by photo-... 

CO2 fixation is also found in many bacteria, both photosynthetic and non-photosynthetic. The purple sulfur and purple nonsulfur bacteria employ the RPP cycle as do plants. The photosynthetic green bacteria, however, use a group of ferre-doxin-linked carboxylases in a pathway known as the reductive carboxylic acid cycle [ ] ... 

The present studies confirm the earlier studies indicating the relatively great biosynthetic abilities of the methane bacteria and suggest that much of the cellular carbon compounds are probably synthesized from acetate and carbon dioxide. In view of the carbon dioxide and acetate requirements and the reductive carboxylation reactions shown to be involved in isoleucine synthesis in M. ruminantium (26) and the probability of similar carboxylation reactions in biosynthesis of isoleucine, alanine, and other amino acids in MOH, suggested by the studies on M. omelianskii (34), the operation of the pyruvate synthase reaction and some other reactions of the reductive carboxylic acid cycle (35, 36) as major pathways of biosynthesis of cellular materials in these bacteria is an attractive hypothesis. 

In the reductive carboxylic acid cycle of photosynthetic bacteria (92-94), which plays a role in the conversion of CO2 to precursors of fatty acids, amino acids, and porphyrins, four molecules of CO2 are fixed in one turn two of them are fixed to a-carbons of carbonyl groups of acetylcoenzyme A and succinyl-coenzyme A to generate pyruvate and a-keto glutarate, respectively ... 

Micrococcus iuteus Mycobacterium smegmatis Rhodopseudomonas viridis (-1- Reductive carboxylic acid cycle) Fru-6-P ... 

R. giobiformis R. sphaeroides Rhodocycius purpureus and reductive carboxylic acid cycle and Fru-1,6-bis-P ... 

Rhodospiriiium rubrum Rhodospiriiium tenue Reductive carboxylic acid cycle VI Pyruvate None... 

FiU. 6. The carbon reduction cycle of photosynthesis. Solid arrows indicate reactions of the carbon reduction cycle as formulated by Calvin and coworkers. Dashed line represents hypothetical reductive carboxylation reaction discussed in text. Open arrows indicate start of some of the biosynthetic paths leading from intermediate compounds of the cycle. Asterisks indicate approximate relative degree of labeling after a few seconds of photosynthesis. They reflect the results of degradation studies by various workers, as discussed in the text. 

These requirements are for the cycle as written in Fig. 6. It has been suggested that in vivo the carboxylation of ribulose-1,5-diphosphate might be a reductive carboxylation (Wilson and Calvin, 1955). Broken isolated chloroplasts and cell-free systems perform only the nonreductive carboxylation of ribulose diphosphate. It is a hypothesis, at present unproved, that in vivo enzyme systems capable of using electrons more directly from the light reactions could catalyze reductive carboxylation [Eq. (23) of Fig. 6]. Such a system might be disrupted when chloroplasts are removed from the cells. In a cycle with a reductive carboxylation, the cofactor requirements might be different. For each complete cycle (three molecules of CO2 taken up) three of the ATP molecules would not be needed if three of the NADPH molecules could be replaced by molecules of reduced ferredoxin. The total requirement would then be six ATP, three NADPH, and six reduced ferredoxin molecules per three CO2 molecules taken up. 

As for the carbon reduction cycle, probably the most important unanswered questions have to do with the mechanisms of the carboxylation reaction and other steps in the cycle. Although all of the biochemical evidence from isolated enzyme systems suggests that the carboxylation reaction is a nonreductive carboxylation of ribulose diphosphate leading to the formation of two molecules of PGA, kinetic evidence with whole cells indicates the possibility of a reductive carboxylation leading to the formation of one molecule of PGA and one molecule of triose phosphate. If this reductive carboxylation does occur, it may be that electrons are somehow conveyed directly from the light reaction to the carbon reduction cycle (Bassham, 1964). If there is such a difference between the... 

In the R.c.c., two of the reactions of the TCA cycle are replaced by alternative reactions catalysed by non-TCA cycle enzymes (i) the citrate (si)-synthase (EC 4.1.3.7)-catalysed formation of citrate from ace-tyl-CoA and oxaloacetate is replaced by the ATP citrate (pro-3.S)-lyase (EC 4.1.3.8)-catalysed, ATP-dri-ven cleavage of citrate to acetyl-CoA and oxaloacetate (eq. 1 J, Fig.), and (ii) the a-ketoglutaiate dehydrogenase complex-catalysed oxidative decarboxylation of a-ketoglutarate to CO2 and succinyl-CoA is replaced by the a-ketoglutarate synthase (EC 1.2.73)-catalysed reductive carboxylation of succinyl CoA (eq. 2 O, Fig.), in which the reductant is reduced ferredoxin (Fd, ... 

Amon and coworkers< > have postulated a new cyclic pathway, named reductive carboxylic acid cycle, that provides another mechanism independent of the reductive pentose cycle for the assimilation of CO2 in bacterial photosynthesis. This cycle generates acetyl-CoA from two molecules of CO2 and is driven by reduced ferredoxin, reduced nucleotides, and ATP, according to the overall simplified equation ... 

Figure 2 State-of-the art hydrocarboxylation of alkenes/alkynes with CO2 (A), an early mechanistic proposal for the reductive carboxylation of ethylene (B), and recent kinetic considerations in a putative catalytic cycle (C).

In all aerobic organisms, FA biosynthesis occurs via two different metabolic pathways (1) tricarboxylic acid cycle (TCA) or Krebs cycle and (2) reductive carboxylation pathway. Albert Szent-Gyorgyi (1893-1986, Hxmgary) discovered FA catalysis during his study (including on vitamin C) on cellular combustion process (TCA cycle) for which he was awarded the Noble Prize in Physiology or Medicine in 1937 (www.nobelprize.org). TCA involves CO2 fixation coupled with the conversion of pyruvate to oxaloacetate, the precursor to malate and fuma-rate (Fig. 8.3). Reductive CO2 fixation catalyzed by the enzyme pyruvate carboxylase xmder... 

Reaction 1 Experimental yield of FA acid in the TCA cycle and reductive carboxylation pathways. 

The fundamental basis of photosynthetic carbon metabolism is the incorporation of carbon dioxide by ribulose-bisphosphate carboxylase (rubisco). This leads to the synthesis of three-carbon sugars which are either exported from the chloroplast or metabolized to regenerate the acceptor ribulose bisphosphate. Rubisco is a bifunctional enzyme in that, in parallel to carboxylation, it catalyzes an oxygenation reaction that leads to phospho-glycolate. This is the starting point for photorespiratory metabolism, which will be discussed below (Section 1.6.2). In C4 plants, the conventional C3 pattern of the photosynthetic carbon reduction Calvin cycle is confined to the bundle sheath cells. The surrounding mesophyll cells act as an ancillary carbon dioxide pump, fixing carbon dioxide via phosphoenolpyruvate carboxylase into C4 acids. These are transported to the bundle sheath for decarboxylation.In this way, photorespiration is limited because of the elevated carbon dioxide levels. 

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