MetaCyc Enzyme: phosphoenolpyruvate carboxylase

Gene: ppc Accession Numbers: EG10756 (MetaCyc), b3956, ECK3947

Synonyms: asp, glu

Species: Escherichia coli K-12 substr. MG1655

Subunit composition of phosphoenolpyruvate carboxylase = [Ppc]4

Phosphoenolpyruvate (PEP) carboxylase (PPC) is a regulatory enzyme, which replenishes oxaloacetate in the tricarboxylic acid cycle [Kameshita79]. It is a unique allosteric enzyme in that it has many kinds of effectors, and has distinct binding sites for each. Cysteinyl, histidyl, lysyl and arginyl residues are essential for catalytic activity. PPC catalyzes the carboxylation of PEP to form oxaloacetate and Pi using Mg2+ or Mn2+ as a cofactor. The reaction proceeds in two steps: formation of carboxyphosphate and the enolate form of pyruvate, followed by carboxylation of the enolate and release of phosphate. [Terada91, Izui83, Kameshita78, Yoshinaga77]

Mutants of the ppc gene have been studied [Terada91, Terada91a, Coomes85, Yano95]. Overexpression of Ppc improves the growth yield on glucose [Chao93] and increases production of succinate from glucose by fermentation [Millard96]. ppc knockout mutants grew slower with less glucose consumption than the parent. The mutants also produced less carbon dioxide and excreted little acetate [Peng04]. Gene expression, enzyme activities, metabolite concentrations and metabolic flux have been measured in a ppc mutant [Peng04, Peng04a]. Site directed mutagenesis of ppc has shown that a conserved histidine residue is essential for the second partial reaction [Terada91].

ppc is essential for growth on glycerol minimal medium [Joyce06].

ppc has a highly conserved sequence in which a GRGG motif is repeated two times with intervening residues. Arg587 is located in this conserved region and was shown to be essential for catalytic activity [Yano95].

The crystal structure for PPC complexed with L-aspartate, Mn2+ or Mn2+ along with a PEP analog and L-aspartate were determined [Kai99, Matsumura99, Matsumura02]. PPC is a tetramer which is made up of a dimer of dimers. Mn2+ and a PEP analog have been shown to directly interact with catalytically essential residues [Kai03].

Extensive studies on the physical and molecular properties of E. coli strain W and B shows PPC to be a tetramer of approximately 400 kDa with monomers around 100 kDa [Yoshinaga70, Smith71, Wohl72].

Citations: [Inoue89, Tan13, Kwon08, Vandedrinck01, Ishijima86, Sabe84, Kodaki84, Canovas65]

Locations: cytosol

Map Position: [4,148,470 <- 4,151,121]

Molecular Weight of Polypeptide: 99.062 kD (from nucleotide sequence)

pI: 5.74 [VanBogelen92]

Unification Links: ASAP:ABE-0012950 , CGSC:368 , DIP:DIP-10538N , EchoBASE:EB0749 , EcoGene:EG10756 , EcoliWiki:b3956 , ModBase:P00864 , OU-Microarray:b3956 , PortEco:ppc , PR:PRO_000023576 , Pride:P00864 , Protein Model Portal:P00864 , RefSeq:NP_418391 , RegulonDB:EG10756 , SMR:P00864 , String:511145.b3956 , UniProt:P00864

Relationship Links: InterPro:IN-FAMILY:IPR015813 , InterPro:IN-FAMILY:IPR018129 , InterPro:IN-FAMILY:IPR021135 , InterPro:IN-FAMILY:IPR022805 , PDB:Structure:1FIY , PDB:Structure:1JQN , PDB:Structure:1QB4 , Pfam:IN-FAMILY:PF00311 , Prints:IN-FAMILY:PR00150 , Prosite:IN-FAMILY:PS00393 , Prosite:IN-FAMILY:PS00781

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

GO Terms:

Biological Process: GO:0006099 - tricarboxylic acid cycle Inferred by computational analysis [GOA01a]
GO:0006107 - oxaloacetate metabolic process Inferred by computational analysis [GOA06]
GO:0008152 - metabolic process Inferred by computational analysis [UniProtGOA11a]
GO:0015977 - carbon fixation Inferred by computational analysis [UniProtGOA11a, GOA06, GOA01a]
Molecular Function: GO:0008964 - phosphoenolpyruvate carboxylase activity Inferred from experiment Inferred by computational analysis [GOA06, GOA01, GOA01a, Izui81]
GO:0000287 - magnesium ion binding Inferred by computational analysis [GOA06]
GO:0003824 - catalytic activity Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0016829 - lyase activity Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Ishihama08, LopezCampistrou05, Terada91]

MultiFun Terms: metabolism energy metabolism, carbon fermentation
metabolism energy metabolism, carbon TCA cycle

Imported from EcoCyc 02-Jun-2015 by Paley S , SRI International

Enzymatic reaction of: phosphoenolpyruvate carboxylase

Synonyms: orthophosphate:oxaloacetate carboxy-lyase (phosphorylating)

EC Number:

oxaloacetate + phosphate <=> phosphoenolpyruvate + hydrogen carbonate

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction in which it was curated.

This reaction is reversible.

Alternative Substrates for phosphoenolpyruvate: phosphoenol-2-oxobutanoate [Terada91 ]

In Pathways: mixed acid fermentation

Imported from EcoCyc 02-Jun-2015 by Paley S , SRI International

Extensive kinetic studies have been performed with E. coli B and W. [Smith80, Kameshita79, Yoshinaga77, Smith73, Wohl72, Silverstein72, Smith71, Izui70]

Cofactors or Prosthetic Groups: Mg2+ [Izui81]

Activators (Allosteric): GTP [Izui81] , fructose 1,6-bisphosphate [Izui81] , acetyl-CoA [Izui81]

Activators (Unknown Mechanism): dioxane [Izui83] , an alcohol [Izui83] , CDP , CMP

Inhibitors (Allosteric): (S)-malate [Izui81] , L-aspartate [Izui81]

Inhibitors (Competitive): phospholactate [Comment 1]

Inhibitors (Unknown Mechanism): (RS)-phosphomalate [Izui83]

Primary Physiological Regulators of Enzyme Activity: GTP , fructose 1,6-bisphosphate , acetyl-CoA , (S)-malate , L-aspartate

Kinetic Parameters:

Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)
[Wohl72, BRENDA14]
[Kai99, BRENDA14]
hydrogen carbonate
[Kai99, BRENDA14]

Sequence Features

Feature Class Location Citations Comment
Active-Site 138
Mutagenesis-Variant 138
[Terada91, UniProt11]
UniProt: Loss of activity.
Active-Site 546
UniProt: Non-Experimental Qualifier: by similarity;
Mutagenesis-Variant 579
[Terada91a, UniProt11]
[Terada91a, UniProt11]
N: 29% of wild-type activity.
P: 5.4% of wild-type activity.
Mutagenesis-Variant 587
[Yano95, UniProt11]
UniProt: Loss of activity.

10/20/97 Gene b3956 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10756; confirmed by SwissProt match.


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

Canovas65: Canovas JL, Kornberg HL (1965). "Fine control of phosphopyruvate carboxylase activity in Escherichia coli." Biochim Biophys Acta 96;169-72. PMID: 14285262

Chao93: Chao YP, Liao JC (1993). "Alteration of growth yield by overexpression of phosphoenolpyruvate carboxylase and phosphoenolpyruvate carboxykinase in Escherichia coli." Appl Environ Microbiol 59(12);4261-5. PMID: 8285716

Coomes85: Coomes MW, Mitchell BK, Beezley A, Smith TE (1985). "Properties of an Escherichia coli mutant deficient in phosphoenolpyruvate carboxylase catalytic activity." J Bacteriol 164(2);646-52. PMID: 3902793

DiazMejia09: Diaz-Mejia JJ, Babu M, Emili A (2009). "Computational and experimental approaches to chart the Escherichia coli cell-envelope-associated proteome and interactome." FEMS Microbiol Rev 33(1);66-97. PMID: 19054114

GOA01: GOA, MGI (2001). "Gene Ontology annotation based on Enzyme Commission mapping." Genomics 74;121-128.

GOA01a: GOA, DDB, FB, MGI, ZFIN (2001). "Gene Ontology annotation through association of InterPro records with GO terms."

GOA06: GOA, SIB (2006). "Electronic Gene Ontology annotations created by transferring manual GO annotations between orthologous microbial proteins."

Inoue89: Inoue M, Hayashi M, Sugimoto M, Harada S, Kai Y, Kasai N, Terada K, Izui K (1989). "First crystallization of a phosphoenolpyruvate carboxylase from Escherichia coli." J Mol Biol 208(3);509-10. PMID: 2677392

Ishihama08: Ishihama Y, Schmidt T, Rappsilber J, Mann M, Hartl FU, Kerner MJ, Frishman D (2008). "Protein abundance profiling of the Escherichia coli cytosol." BMC Genomics 9;102. PMID: 18304323

Ishijima86: Ishijima S, Izui K, Katsuki H (1986). "Phosphoenolpyruvate carboxylase of Escherichia coli K-12. N- and C-terminal sequences and tentative assignment of the catalytically essential cysteine residue." J Biochem 99(5);1299-310. PMID: 3519602

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

Joyce06: Joyce AR, Reed JL, White A, Edwards R, Osterman A, Baba T, Mori H, Lesely SA, Palsson BO, Agarwalla S (2006). "Experimental and computational assessment of conditionally essential genes in Escherichia coli." J Bacteriol 188(23);8259-71. PMID: 17012394

Kai03: Kai Y, Matsumura H, Izui K (2003). "Phosphoenolpyruvate carboxylase: three-dimensional structure and molecular mechanisms." Arch Biochem Biophys 414(2);170-9. PMID: 12781768

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

Kameshita78: Kameshita I, Tokushige M, Katsuki H (1978). "Phosphoenolpyruvate carboxylase of Escherichia coli. Essential arginyl residues for catalytic and regulatory functions." J Biochem (Tokyo) 1978;84(4);795-803. PMID: 361729

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

Kodaki84: Kodaki T, Fujita N, Kameshita I, Izui K, Katsuki H (1984). "Phosphoenolpyruvate carboxylase of Escherichia coli. Specificity of some compounds as activators at the site for fructose 1,6-bisphosphate, one of the allosteric effectors." J Biochem 95(3);637-42. PMID: 6373747

Kwon08: Kwon YD, Lee SY, Kim P (2008). "A physiology study of Escherichia coli overexpressing phosphoenolpyruvate carboxykinase." Biosci Biotechnol Biochem 72(4);1138-41. PMID: 18391462

LopezCampistrou05: Lopez-Campistrous A, Semchuk P, Burke L, Palmer-Stone T, Brokx SJ, Broderick G, Bottorff D, Bolch S, Weiner JH, Ellison MJ (2005). "Localization, annotation, and comparison of the Escherichia coli K-12 proteome under two states of growth." Mol Cell Proteomics 4(8);1205-9. PMID: 15911532

Matsumura02: Matsumura H, Xie Y, Shirakata S, Inoue T, Yoshinaga T, Ueno Y, Izui K, Kai Y (2002). "Crystal structures of C4 form maize and quaternary complex of E. coli phosphoenolpyruvate carboxylases." Structure 10(12);1721-30. PMID: 12467579

Matsumura99: Matsumura H, Terada M, Shirakata S, Inoue T, Yoshinaga T, Izui K, Kai Y (1999). "Plausible phosphoenolpyruvate binding site revealed by 2.6 A structure of Mn2+-bound phosphoenolpyruvate carboxylase from Escherichia coli." FEBS Lett 458(2);93-6. PMID: 10481043

Millard96: Millard CS, Chao YP, Liao JC, Donnelly MI (1996). "Enhanced production of succinic acid by overexpression of phosphoenolpyruvate carboxylase in Escherichia coli." Appl Environ Microbiol 62(5);1808-10. PMID: 8633880

Peng04: Peng L, Shimizu K (2004). "Effect of ppc gene knockout on the metabolism of Escherichia coli in view of gene expressions, enzyme activities and intracellular metabolite concentrations." Appl Microbiol Biotechnol NIL. PMID: 14963616

Peng04a: Peng L, Arauzo-Bravo MJ, Shimizu K (2004). "Metabolic flux analysis for a ppc mutant Escherichia coli based on 13C-labelling experiments together with enzyme activity assays and intracellular metabolite measurements." FEMS Microbiol Lett 235(1);17-23. PMID: 15158257

Sabe84: Sabe H, Miwa T, Kodaki T, Izui K, Hiraga S, Katsuki H (1984). "Molecular cloning of the phosphoenolpyruvate carboxylase gene, ppc, of Escherichia coli." Gene 31(1-3);279-83. PMID: 6396163

Silverstein72: Silverstein R (1972). "Kinetic studies of acetyl coenzyme A activated phosphoenolpyruvate carboxylase: reverse effects with a fatty acid." Biochim Biophys Acta 258(2);626-36. PMID: 4551567

Smith71: Smith TE (1971). "Escherichia coli phosphoenolpyruvate carboxylase. Physical and chemical properties." J Biol Chem 246(13);4234-41. PMID: 4932977

Smith73: Smith TE, Perry M (1973). "Escherichia coli phosphoenolpyruvate carboxylase: kinetics and mechanism of inactivation." Arch Biochem Biophys 156(2);448-55. PMID: 4578117

Smith80: Smith TE, Balasubramanian KA, Beezley A (1980). "Escherichia coli phosphoenolpyruvate carboxylase. Studies on the mechanism of synergistic activation by nucleotides." J Biol Chem 255(4);1635-42. PMID: 6986377

Tan13: Tan Z, Zhu X, Chen J, Li Q, Zhang X (2013). "Activating phosphoenolpyruvate carboxylase and phosphoenolpyruvate carboxykinase in combination for improvement of succinate production." Appl Environ Microbiol 79(16);4838-44. PMID: 23747698

Terada91: Terada K, Izui K (1991). "Site-directed mutagenesis of the conserved histidine residue of phosphoenolpyruvate carboxylase. His138 is essential for the second partial reaction." Eur J Biochem 1991;202(3);797-803. PMID: 1765093

Terada91a: Terada K, Murata T, Izui K (1991). "Site-directed mutagenesis of phosphoenolpyruvate carboxylase from E. coli: the role of His579 in the catalytic and regulatory functions." J Biochem (Tokyo) 109(1);49-54. PMID: 2016273

UniProt10a: UniProt Consortium (2010). "UniProt version 2010-07 released on 2010-06-15 00:00:00." Database.

UniProt11: UniProt Consortium (2011). "UniProt version 2011-06 released on 2011-06-30 00:00:00." Database.

UniProt15: UniProt Consortium (2015). "UniProt version 2015-01 released on 2015-01-16 00:00:00." Database.

UniProtGOA11a: UniProt-GOA (2011). "Gene Ontology annotation based on manual assignment of UniProtKB keywords in UniProtKB/Swiss-Prot entries."

VanBogelen92: VanBogelen RA, Sankar P, Clark RL, Bogan JA, Neidhardt FC (1992). "The gene-protein database of Escherichia coli: edition 5." Electrophoresis 1992;13(12);1014-54. PMID: 1286664

Vandedrinck01: Vandedrinck S, Deschamps G, Sablon E, Vandamme EJ (2001). "Construction and characterization of a PPC (phosphoenolpyruvate carboxylase) knockout mutant of Escherichia coli." Meded Rijksuniv Gent Fak Landbouwkd Toegep Biol Wet 66(4);345-9. PMID: 15954317

Wohl72: Wohl RC, Markus G (1972). "Phosphoenolpyruvate carboxylase of Escherichia coli. Purification and some properties." J Biol Chem 247(18);5785-92. PMID: 4560418

Yano95: Yano M, Terada K, Umiji K, Izui K (1995). "Catalytic role of an arginine residue in the highly conserved and unique sequence of phosphoenolpyruvate carboxylase." J Biochem (Tokyo) 117(6);1196-200. PMID: 7490260

Yoshinaga70: Yoshinaga T, Izui K, Katsuki H (1970). "Purification and molecular properties of allosteric phosphoenolpyruvate carboxylase from Escherichia coli." J Biochem 68(5);747-50. PMID: 4921962

Yoshinaga77: Yoshinaga T (1977). "Structural specificity of the allosteric inhibitor of phosphoenolpyruvate carboxylase of Escherichia coli." J Biochem (Tokyo) 1977;81(3);665-71. PMID: 324991

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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
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