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MetaCyc Pathway: C4 photosynthetic carbon assimilation cycle, NADP-ME type
Inferred from experiment

Pathway diagram: C4 photosynthetic carbon assimilation cycle, NADP-ME type

This view shows enzymes only for those organisms listed below, in the list of taxa known to possess the pathway. If an enzyme name is shown in bold, there is experimental evidence for this enzymatic activity.

Synonyms: PCA cycle, C4 photosynthesis, NADP-ME type

Superclasses: Generation of Precursor Metabolites and EnergyPhotosynthesis

Some taxa known to possess this pathway include : Zea mays

Expected Taxonomic Range: Embryophyta

General Background

The main CO2 fixation enzyme in plants, EC, ribulose-bisphosphate carboxylase/oxygenase (RuBisCO), has both carboxylase (photosynthetic CO2 fixation) and oxygenase (photorespiration) activities. The carboxylase activity is favored at high CO2/O2 ratio, whereas the oxygenase activity predominates at a decreased CO2/O2 ratio.

At the time when RuBisCO first evolved, atmospheric CO2 concentration was very high and thus the enzyme's carboxylase activity was favored. However, later in evolution, atmospheric O2 concentration began to rise and RuBisCO's oxygenase activity increased at the expense of the carboxylase activity. At current CO2 levels, photorespiration can reduce photosynthesis by more than 40%.

C4 plants have evolved to respond to lowered CO2 levels. The C4 photosynthetic carbon assimilation cycle starts with carbon fixation in the mesophyll cells by a combination of EC, carbonic anhydrase, which converts the gasous CO2 to soluble hydrogen carbonate, and EC, phosphoenolpyruvate carboxylase, which incorporates the latter, forming the C4 acid oxaloacetate. Oxaloacetate or a derivative thereof ( L-aspartate) is then transported from the mesophyll cells to bundle sheath cells. Once inside the bundle sheath cells, CO2 is released from the C4 acids, generating a micro-environment with a much higher concentration of CO2, suitable for an efficient carboxylation by RuBisCO.

There are at least three variations of the C4 photosynthetic carbon assimilation cycle in plants, namely the NADP-ME (NADP-malic enzyme), NAD-ME (NAD-malic enzyme), or PEPCK (phosphoenolpyruvate carboxykinase) types. The NADP-ME variant does not involve L-aspartate and utilizes an NADP-dependent malic enzyme to releases CO2 from (S)-malate. The NAD-ME variant does involve L-aspartate and utilizes an NAD-dependent malic enzyme. The PEPCK variant contains an additional route for CO2 release by EC, phosphoenolpyruvate carboxykinase (ATP), which usually operates to supplement the NADP-ME variant.

However, recent research indicates a flexibility between the three subtypes of C4 photosynthesis (see also C4 photosynthetic carbon assimilation cycle, NAD-ME type and C4 photosynthetic carbon assimilation cycle, PEPCK type) which is documented by the occurrence of mixed C4 decarboxylation pathways in carious plants previously considered to belong to exclusively one C4 photosynthetic type [Wang14b]. For instance, Zea mays has been shown to use both L-aspartate and oxaloacetate for the generation of CO2 [Chapman81]. In addition, it has been demonstrated that maize leaves exhibit the corresponding aspartate aminotransferase activity to maintain an efficient flow-rate. Moreover, the phosphoenol carboxykinase (PEPCK), considered the hallmark of the PEPCK pathway, is also active in maize and shown to support L-aspartate-driven photosynthesis [Pick11]. Similar results have been reported from the NADP-ME type plant Flaveria bidentis which also utilized both L-aspartate and oxaloacetate equally and demonstrated aspartate aminotransferase activity [Meister96]. In general, current discussion of the C4 photosynthetic pathway types move in a direction favoring the existence of only two subtypes, i.e. NADP-ME and NAD-ME which may both contain the supplementary PEPCK cycle [Wang14b].

About This Pathway

C4 photosynthesis basically represents a CO2 concentrating mechanism (CCM) which independently originated more than 60 times in plants indicating the evolutionary drive to optimize photosynthetic CO2 fixation as an tool to keep pace with an ever-changing environment [Ludwig12].This MetaCyc pathway describes the NADP-ME variant, found in major C4 crops such as maize, sorghum, and sugar cane. Note that the pathway spans two cell types, mesophyll and bundle sheath cells.

The roles and sub-cellular locations of carbonic anhydrases (CA) in plants differ and are metabolically integrated in a rather complex manner regardless of the seemingly simple reaction they catalyze, i.e. the reversible hydration of CO2 to hydrogen carbonate [Badger03, Moroney01]. In C3 plants CA's are ambivalent enzymes and contribute to increased CO2 concentration in micro-compartments creating a more efficient environment for the carbon fixating role of RuBisCO in the chloroplast, and also have non-photosynthetic functions as in lipid biosynthesis and operating as antioxidant [Rowlett14]. However, in C4 plants β-carbonic anhydrases operating in the cytoplasm of mesophyll cells are considered essential for the operation of the pathway by providing the PEP-carboxylase substrate hydrogen carbonate for CO2 fixation into oxaloacetate [Burnell88]. Corresponding β-carbonic anhydrases characterized in Zea mays, i.e. CA2 [Tems10, Burnell97] and in Flaveria bidentis [Tetu07], i.e. CA3 are part of a small CA family in this plants where only one member is directly involved in the first reaction of C4 photosynthesis [Monti13, Burnell90]. However, it appears that even those β-carbonic anhydrases have only a limited role in C4 photosynthesis under current climate conditions and are more essential when C4 plants are stressed by high temperature and drought [Studer14].

The initial CO2 fixation occurs in the mesophyll cells, producing oxaloacetate that is subsequently converted to (S)-malate. Malate is transported into bundle sheath cells and CO2 is released there by an NADP-dependent malic enzyme. Pyruvate reenters the mesophyll cells and replenishes phosphoenolpyruvate.

Experimental evidence indicates that CO2 fixation in cyanobacteria is mediated by the C3 pathway, and although C4 acids are among the initial products of photosynthesis, these organisms do not have the enzymatic capacity for the operation of a C4 pathway [Colman89].

Revised 05-Sep-2014 by Caspi R, SRI International
Revised 24-Jan-2015 by Foerster H, Boyce Thompson Institute


Badger03: Badger M (2003). "The roles of carbonic anhydrases in photosynthetic CO2 concentrating mechanisms." Photosynthesis Research 77:83-94.

Beverly89: Beverly Rothermel, Timothy Nelson J. Bio. Chem. (1989) 264: 19587-19592.

Burnell88: Burnell JN, Hatch MD (1988). "Low bundle sheath carbonic anhydrase is apparently essential for effective c(4) pathway operation." Plant Physiol 86(4);1252-6. PMID: 16666063

Burnell90: Burnell JN (1990). "Immunological Study of Carbonic Anhydrase in C3 and C4 Plants Using Antibodies to Maize Cytosolic and Spinach Chloroplastic Carbonic Anhydrase." Plant Cell Physiol. 31(4):423-427.

Burnell97: Burnell JN, Ludwig M (1997). "Characterisation of Two cDNAs Encoding Carbonic Anhydrase in Maize Leaves." Australian Journal of Plant Physiology 24(4):451-458.

Chapman81: Chapman KSR, Hatch MD (1981). "Aspartate Decarboxylation in Bundle Sheath Cells of Zea mays and its possible Contribution to C4 Photosynthesis." Australian Journal of Plant Physiology 8(2):237-248.

Colman89: Colman, B. (1989). "Photosynthetic Carbon Assimilation and the Suppression of Photorespiration in the Cyanobacteria." quatic Botany 34:211-231.

Ludwig12: Ludwig M (2012). "Carbonic anhydrase and the molecular evolution of C4 photosynthesis." Plant Cell Environ 35(1);22-37. PMID: 21631531

Meister96: Meister M, Agostino A, Hatch MD (1996). "The roles of malate and aspartate in C4 photosynthetic metabolism of Flaveria bidentis (L.)." Planta 199:262-269.

Monti13: Monti SM, De Simone G, Dathan NA, Ludwig M, Vullo D, Scozzafava A, Capasso C, Supuran CT (2013). "Kinetic and anion inhibition studies of a β-carbonic anhydrase (FbiCA 1) from the C4 plant Flaveria bidentis." Bioorg Med Chem Lett 23(6);1626-30. PMID: 23414801

Moroney01: Moroney JV, Bartlett SG, Samuelsson G (2001). "Carbonic anhydrase in plants and algae." Plant, Cell & Environment 24(2):141-153.

Pick11: Pick TR, Brautigam A, Schluter U, Denton AK, Colmsee C, Scholz U, Fahnenstich H, Pieruschka R, Rascher U, Sonnewald U, Weber AP (2011). "Systems analysis of a maize leaf developmental gradient redefines the current C4 model and provides candidates for regulation." Plant Cell 23(12);4208-20. PMID: 22186372

Rowlett14: Rowlett RS (2014). "Structure and catalytic mechanism of β-carbonic anhydrases." Subcell Biochem 75;53-76. PMID: 24146374

Studer14: Studer AJ, Gandin A, Kolbe AR, Wang L, Cousins AB, Brutnell TP (2014). "A Limited Role for Carbonic Anhydrase in C4 Photosynthesis as Revealed by a ca1ca2 Double Mutant in Maize." Plant Physiol 165(2);608-617. PMID: 24706552

Tems10: Tems U, Burnell JN (2010). "Characterization and expression of the maize β-carbonic anhydrase gene repeat regions." Plant Physiol Biochem 48(12);945-51. PMID: 20933433

Tetu07: Tetu SG, Tanz SK, Vella N, Burnell JN, Ludwig M (2007). "The Flaveria bidentis beta-carbonic anhydrase gene family encodes cytosolic and chloroplastic isoforms demonstrating distinct organ-specific expression patterns." Plant Physiol 144(3);1316-27. PMID: 17496111

Wang14b: Wang Y, Brautigam A, Weber AP, Zhu XG (2014). "Three distinct biochemical subtypes of C4 photosynthesis? A modelling analysis." J Exp Bot 65(13);3567-78. PMID: 24609651

Other References Related to Enzymes, Genes, Subpathways, and Substrates of this Pathway

Bologna07: Bologna FP, Andreo CS, Drincovich MF (2007). "Escherichia coli malic enzymes: two isoforms with substantial differences in kinetic properties, metabolic regulation, and structure." J Bacteriol 189(16);5937-46. PMID: 17557829

BRENDA14: BRENDA team (2014). Imported from BRENDA version existing on Aug 2014.

Brown81: Brown DA, Cook RA (1981). "Role of metal cofactors in enzyme regulation. Differences in the regulatory properties of the Escherichia coli nicotinamide adenine dinucleotide phosphate specific malic enzyme, depending on whether magnesium ion or manganese ion serves as divalent cation." Biochemistry 1981;20(9);2503-12. PMID: 7016178

Cavallaro94: Cavallaro A, Ludwig M, Burnell J (1994). "The nucleotide sequence of a complementary DNA encoding Flaveria bidentis carbonic anhydrase." FEBS Lett 350(2-3);216-8. PMID: 8070567

Ferte86: Ferte N, Jacquot JP, Meunier JC (1986). "Structural, immunological and kinetic comparisons of NADP-dependent malate dehydrogenases from spinach (C3) and corn (C4) chloroplasts." Eur J Biochem 154(3);587-95. PMID: 3948869

Fett94: Fett JP, Coleman JR (1994). "Characterization and expression of two cDNAs encoding carbonic anhydrase in Arabidopsis thaliana." Plant Physiol 105(2);707-13. PMID: 7520589

Furumoto00: Furumoto, T et. al. Plant Cell Physiol (2000) 41:1200-1209


Gennidakis07: Gennidakis S, Rao S, Greenham K, Uhrig RG, O'Leary B, Snedden WA, Lu C, Plaxton WC (2007). "Bacterial- and plant-type phosphoenolpyruvate carboxylase polypeptides interact in the hetero-oligomeric Class-2 PEPC complex of developing castor oil seeds." Plant J 52(5);839-49. PMID: 17894783

Guilloton92: Guilloton MB, Korte JJ, Lamblin AF, Fuchs JA, Anderson PM (1992). "Carbonic anhydrase in Escherichia coli. A product of the cyn operon." J Biol Chem 1992;267(6);3731-4. PMID: 1740425

Izui70: Izui K (1970). "Kinetic studies on the allosteric nature of phosphoenolpyruvate carboxylase from Escherichia coli." J Biochem 68(2);227-38. PMID: 4917116

Izui81: Izui K, Taguchi M, Morikawa M, Katsuki H (1981). "Regulation of Escherichia coli phosphoenolpyruvate carboxylase by multiple effectors in vivo. II. Kinetic studies with a reaction system containing physiological concentrations of ligands." J Biochem 90(5);1321-31. PMID: 7040354

Izui83: Izui K, Matsuda Y, Kameshita I, Katsuki H, Woods AE (1983). "Phosphoenolpyruvate carboxylase of Escherichia coli. Inhibition by various analogs and homologs of phosphoenolpyruvate." J Biochem (Tokyo) 1983;94(6);1789-95. PMID: 6368527

Kagawa88: Kagawa T, Bruno PL (1988). "NADP-malate dehydrogenase from leaves of Zea mays: purification and physical, chemical, and kinetic properties." Arch Biochem Biophys 260(2);674-95. PMID: 3341761

Kai99: Kai Y, Matsumura H, Inoue T, Terada K, Nagara Y, Yoshinaga T, Kihara A, Tsumura K, Izui K (1999). "Three-dimensional structure of phosphoenolpyruvate carboxylase: a proposed mechanism for allosteric inhibition." Proc Natl Acad Sci U S A 96(3);823-8. PMID: 9927652

Kameshita79: Kameshita I, Tokushige M, Izui K, Katsuki H (1979). "Phosphoenolpyruvate carboxylase of Escherichia coli. Affinity labeling with bromopyruvate." J Biochem (Tokyo) 1979;86(5);1251-7. PMID: 42643

Keilin39: Keilin, D., Mann, T. (1939). "Carbonic anhydrase." Nature 144:442-443.

Krall93: Krall, John, Edwards, Gerald (1993). "PEP Carboxylases from Two C4 Species of Panicum with Markedly DifferentSusceptibilities to Cold Inactivation." Plant Cell Physiol. 34(1): 1-11.

Latendresse13: Latendresse M. (2013). "Computing Gibbs Free Energy of Compounds and Reactions in MetaCyc."

Matsumura99: Matsumura H, Nagata T, Terada M, Shirakata S, Inoue T, Yoshinaga T, Ueno Y, Saze H, Izui K, Kai Y (1999). "Crystallization and preliminary x-ray diffraction studies of C4-form phosphoenolpyruvate carboxylase from maize." Acta Crystallogr D Biol Crystallogr 1999;55(11);1937-8. PMID: 10531501

Matsuoka88a: Matsuoka M, Ozeki Y, Yamamoto N, Hirano H, Kano-Murakami Y, Tanaka Y (1988). "Primary structure of maize pyruvate, orthophosphate dikinase as deduced from cDNA sequence." J Biol Chem 263(23);11080-3. PMID: 2841317

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Please cite the following article in publications resulting from the use of MetaCyc: Caspi et al, Nucleic Acids Research 42:D459-D471 2014
Page generated by Pathway Tools version 19.5 (software by SRI International) on Mon May 2, 2016, biocyc14.