|Gene:||gapA||Accession Numbers: EG10367 (MetaCyc), b1779, ECK1777|
Synonyms: gad, gap1, GAPDH-A
Species: Escherichia coli K-12 substr. MG1655
Glyceraldehyde 3-phosphate dehydrogenase A catalyzes the reversible oxidative phosphorylation of D-glyceraldehyde-3-phosphate to 1,3-bisphospho-D-glycerate in the presence of NAD+ and phosphate during glycolysis and gluconeogenesis in E. coli. The enzyme is also found in many other organisms and its properties have been extensively studied (see [ENZYMESVOLXIII76]).
E. coli is unusual in having two glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activities encoded by gapA and epd (gapB). However, the gapA encoded enzyme has a highly efficient phosphorylating glyceraldehyde-3-phosphate dehydrogenase activity and a low phosphorylating erythrose-4-phosphate dehydrogenase activity, whereas the epd encoded enzyme has an efficient non-phosphorylating erythrose-4-phosphate dehydrogenase activity and a very low phosphorylating glyceraldehyde-3-phosphate dehydrogenase activity [Zhao95, BoschiMuller97].
The GapA protein has a sequence that is more similar to eukaryotic sequences than to the thermophilic bacterial enzymes, and to prokaryotic enzymes in general [Doolittle90, Branlant85]. The gapA product is required for glycolysis, while the epd product is not [Seta97]. Both enzymes may be involved in production of pyridoxal 5'-phosphate (PLP) [Yang98a].
Early studies of gapA mutants from E. coli K-10 implicated its role in glycolysis and demonstrated some of its catalytic properties [Hillman75, Hillman79]. A gapA mutant exhibits a growth defect and also exhibits increased aggregation and lysis phenotypes that are rescued by high-salt media [Seta97].
Regulation of gapA gene expression has been studied [Charpentier94, Charpentier98, Riehle03, Thouvenot04]. The regulation of the fkpA, gapA, and hslT genes is affected by evolution under conditions of chronic heat stress [Riehle03].
The crystal structure of the wild-type enzyme in the presence of NAD+ has been determined at 1.80 Å resolution and was similar to those of other GAPDHs. The crystal structure of a N313T mutant was also determined at 2.17 Å resolution [Duee96]. Several other E. coli GAPDH crystal structures have been reported with and without bound NAD+, and in the hemiacetal intermediate state [Yun00].
Molecular factors responsible for the NAD+ cofactor stereospecificity have been studied using site-directed mutagenesis. The enzyme is a B-specific dehydrogenase that catalyzes transfer of the pro-S hydrogen and binds NAD(H) in the syn nicotinamide orientation [Eyschen99]. Refolding of denatured E. coli GAPDH in the presence of chaperone protein Tig; trigger factor has been studied [Huang00].
ADP-ribosylated GAPDH is a secreted virulence factor in some fungi and Gram-positive pathogens, as well as in pathogenic strains of E. coli. Non-pathogenic E. coli do not secrete GAPDH [Egea07, Aguilera09]. Evidence suggests that E. coli GAPDH is also involved in DNA repair [Ferreira13, Ferreira15].
A series of vectors inducibly expressing paired-terminus antisense RNAs was constructed to silence central carbon metabolism in host E. coli K-12 MG1655. A vector that silenced gapA at 93% efficacy caused severe growth inhibition [Nakashima14]. Regulating the expression of an engineered E. coli gapA through changes in temperature has been demonstrated to control glycolysis [Cho12].
Locations: membrane, cytosol
|Map Position: [1,860,795 -> 1,861,790]|
Molecular Weight of Polypeptide: 35.532 kD (from nucleotide sequence), 35.0 kD (experimental) [DAlessio71a ]
Molecular Weight of Multimer: 144.0 kD (experimental) [DAlessio71]
Unification Links: ASAP:ABE-0005920 , DIP:DIP-31848N , EchoBASE:EB0362 , EcoGene:EG10367 , EcoliWiki:b1779 , Mint:MINT-1255410 , OU-Microarray:b1779 , PortEco:gapA , PR:PRO_000022749 , Pride:P0A9B2 , Protein Model Portal:P0A9B2 , RefSeq:NP_416293 , RegulonDB:EG10367 , SMR:P0A9B2 , String:511145.b1779 , UniProt:P0A9B2
Relationship Links: InterPro:IN-FAMILY:IPR006424 , InterPro:IN-FAMILY:IPR016040 , InterPro:IN-FAMILY:IPR020828 , InterPro:IN-FAMILY:IPR020829 , InterPro:IN-FAMILY:IPR020830 , InterPro:IN-FAMILY:IPR020831 , Panther:IN-FAMILY:PTHR10836 , PDB:Structure:1DC3 , PDB:Structure:1DC4 , PDB:Structure:1DC5 , PDB:Structure:1DC6 , PDB:Structure:1GAD , PDB:Structure:1GAE , PDB:Structure:1S7C , PDB:Structure:2VYN , PDB:Structure:2VYV , Pfam:IN-FAMILY:PF00044 , Pfam:IN-FAMILY:PF02800 , Prints:IN-FAMILY:PR00078 , Prosite:IN-FAMILY:PS00071 , Smart:IN-FAMILY:SM00846
|Biological Process:||GO:0006096 - glycolytic process
[UniProtGOA12, UniProtGOA11, Seta97]
GO:0006006 - glucose metabolic process [GOA01a]
GO:0055114 - oxidation-reduction process [UniProtGOA11, GOA01a]
|Molecular Function:||GO:0004365 - glyceraldehyde-3-phosphate dehydrogenase (NAD+) (phosphorylating) activity
GO:0051287 - NAD binding [GOA01a, Eyschen99]
GO:0016491 - oxidoreductase activity [UniProtGOA11]
GO:0016620 - oxidoreductase activity, acting on the aldehyde or oxo group of donors, NAD or NADP as acceptor [GOA01a]
GO:0050661 - NADP binding [GOA01a]
|Cellular Component:||GO:0005829 - cytosol
[DiazMejia09, Ishihama08, Zhang07, LopezCampistrou05, Bairoch93, Lasserre06, Branlant85]
GO:0016020 - membrane [Lasserre06]
GO:0005737 - cytoplasm [UniProtGOA11a, UniProtGOA11]
|MultiFun Terms:||metabolism → biosynthesis of building blocks → cofactors, small molecule carriers → pyridoxal 5'phosphate|
|metabolism → central intermediary metabolism|
|metabolism → energy metabolism, carbon → glycolysis|
Enzymatic reaction of: glyceraldehyde 3-phosphate dehydrogenase
Synonyms: GAPDH, triosephosphate dehydrogenase
EC Number: 220.127.116.11
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. [Soukri89]
In Pathways: superpathway of hexitol degradation (bacteria) , superpathway of N-acetylneuraminate degradation , superpathway of glycolysis and Entner-Doudoroff , superpathway of glycolysis, pyruvate dehydrogenase, TCA, and glyoxylate bypass , gluconeogenesis I , glycolysis II (from fructose 6-phosphate) , glycolysis I (from glucose 6-phosphate)
The pH optimum was 8.8 in sodium pyrophosphate buffer, and 8.0 in Tris-chloride or triethanolamine-chloride buffer [DAlessio71a]. The enzyme shows esterase activity with p-nitrophenyl acetate as substrate. The esterase activity is inhibited by AMP, ADP and ATP [DAlessio71a].
The enzyme is inhibited by various dihydropyrazine derivatives and the inhibition can be reversed by several sulfhydryl compounds [Takechi10].
Cofactor Binding Comment: GAPDH has a high affinity for its cofactor NAD+. NAD+ is an essential requirement for both catalytic functions (phosphorylative activity and oxidative activity) Although NAD+ is not itself a reactant, its binding apparently effects a conformational change necessary for activity of the phosphorylative head.[Hillman79]
pH(opt): 8.8 [BRENDA14]
|Chain||2 -> 331|
|Nucleotide-Phosphate-Binding-Region||12 -> 13|
|Protein-Segment||149 -> 151|
|Protein-Segment||209 -> 210|
10/20/97 Gene b1779 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10367; confirmed by SwissProt match.
Aguilera09: Aguilera L, Gimenez R, Badia J, Aguilar J, Baldoma L (2009). "NAD+-dependent post-translational modification of Escherichia coli glyceraldehyde-3-phosphate dehydrogenase." Int Microbiol 12(3);187-92. PMID: 19784925
BoschiMuller97: Boschi-Muller S, Azza S, Pollastro D, Corbier C, Branlant G (1997). "Comparative enzymatic properties of GapB-encoded erythrose-4-phosphate dehydrogenase of Escherichia coli and phosphorylating glyceraldehyde-3-phosphate dehydrogenase." J Biol Chem 272(24);15106-12. PMID: 9182530
Branlant85: Branlant G, Branlant C (1985). "Nucleotide sequence of the Escherichia coli gap gene. Different evolutionary behavior of the NAD+-binding domain and of the catalytic domain of D-glyceraldehyde-3-phosphate dehydrogenase." Eur J Biochem 1985;150(1);61-6. PMID: 2990926
Charpentier94: Charpentier B, Branlant C (1994). "The Escherichia coli gapA gene is transcribed by the vegetative RNA polymerase holoenzyme E sigma 70 and by the heat shock RNA polymerase E sigma 32." J Bacteriol 176(3);830-9. PMID: 8300536
Charpentier98: Charpentier B, Bardey V, Robas N, Branlant C (1998). "The EIIGlc protein is involved in glucose-mediated activation of Escherichia coli gapA and gapB-pgk transcription." J Bacteriol 180(24);6476-83. PMID: 9851989
Cho12: Cho HS, Seo SW, Kim YM, Jung GY, Park JM (2012). "Engineering glyceraldehyde-3-phosphate dehydrogenase for switching control of glycolysis in Escherichia coli." Biotechnol Bioeng 109(10);2612-9. PMID: 22528318
DAlessio71: D'Alessio G, Josse J (1971). "Glyceraldehyde phosphate dehydrogenase, phosphoglycerate kinase, and phosphoglyceromutase of Escherichia coli. Simultaneous purification and physical properties." J Biol Chem 1971;246(13);4319-25. PMID: 4932978
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
Duee96: Duee E, Olivier-Deyris L, Fanchon E, Corbier C, Branlant G, Dideberg O (1996). "Comparison of the structures of wild-type and a N313T mutant of Escherichia coli glyceraldehyde 3-phosphate dehydrogenases: implication for NAD binding and cooperativity." J Mol Biol 257(4);814-38. PMID: 8636984
Egea07: Egea L, Aguilera L, Gimenez R, Sorolla MA, Aguilar J, Badia J, Baldoma L (2007). "Role of secreted glyceraldehyde-3-phosphate dehydrogenase in the infection mechanism of enterohemorrhagic and enteropathogenic Escherichia coli: interaction of the extracellular enzyme with human plasminogen and fibrinogen." Int J Biochem Cell Biol 39(6);1190-203. PMID: 17449317
Eyschen99: Eyschen J, Vitoux B, Marraud M, Cung MT, Branlant G (1999). "Engineered glycolytic glyceraldehyde-3-phosphate dehydrogenase binds the anti conformation of NAD+ nicotinamide but does not experience A-specific hydride transfer." Arch Biochem Biophys 364(2);219-27. PMID: 10190977
Ferreira13: Ferreira E, Gimenez R, Aguilera L, Guzman K, Aguilar J, Badia J, Baldoma L "Protein interaction studies point to new functions for Escherichia coli glyceraldehyde-3-phosphate dehydrogenase." Res Microbiol 164(2);145-54. PMID: 23195894
Ferreira15: Ferreira E, Gimenez R, Canas MA, Aguilera L, Aguilar J, Badia J, Baldoma L (2015). "Glyceraldehyde-3-phosphate dehydrogenase is required for efficient repair of cytotoxic DNA lesions in Escherichia coli." Int J Biochem Cell Biol 60;202-12. PMID: 25603270
Lasserre06: Lasserre JP, Beyne E, Pyndiah S, Lapaillerie D, Claverol S, Bonneu M (2006). "A complexomic study of Escherichia coli using two-dimensional blue native/SDS polyacrylamide gel electrophoresis." Electrophoresis 27(16);3306-21. PMID: 16858726
Link97: Link AJ, Robison K, Church GM (1997). "Comparing the predicted and observed properties of proteins encoded in the genome of Escherichia coli K-12." Electrophoresis 18(8);1259-313. PMID: 9298646
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
Nakashima14: Nakashima N, Ohno S, Yoshikawa K, Shimizu H, Tamura T (2014). "A vector library for silencing central carbon metabolism genes with antisense RNAs in Escherichia coli." Appl Environ Microbiol 80(2);564-73. PMID: 24212579
Pasquali94: Pasquali C., Sanchez J.-C., Ravier F., Golaz O., Hughes G.J., Frutiger S., Paquet N., Wilkins M., Appel R.D., Bairoch A., Hochstrasser D.F. (1994). Data submission to UniProtKB on 1994-09.
Peng11: Peng C, Lu Z, Xie Z, Cheng Z, Chen Y, Tan M, Luo H, Zhang Y, He W, Yang K, Zwaans BM, Tishkoff D, Ho L, Lombard D, He TC, Dai J, Verdin E, Ye Y, Zhao Y (2011). "The first identification of lysine malonylation substrates and its regulatory enzyme." Mol Cell Proteomics 10(12);M111.012658. PMID: 21908771
Riehle03: Riehle MM, Bennett AF, Lenski RE, Long AD (2003). "Evolutionary changes in heat-inducible gene expression in lines of Escherichia coli adapted to high temperature." Physiol Genomics 14(1);47-58. PMID: 12672900
Soukri89: Soukri A, Mougin A, Corbier C, Wonacott A, Branlant C, Branlant G (1989). "Role of the histidine 176 residue in glyceraldehyde-3-phosphate dehydrogenase as probed by site-directed mutagenesis." Biochemistry 28(6);2586-92. PMID: 2659073
Thouvenot04: Thouvenot B, Charpentier B, Branlant C (2004). "The strong efficiency of the Escherichia coli gapA P1 promoter depends on a complex combination of functional determinants." Biochem J 383(Pt 2);371-82. PMID: 15250823
Yang98a: Yang Y, Zhao G, Man TK, Winkler ME (1998). "Involvement of the gapA- and epd (gapB)-encoded dehydrogenases in pyridoxal 5'-phosphate coenzyme biosynthesis in Escherichia coli K-12." J Bacteriol 1998;180(16);4294-9. PMID: 9696782
Yun00: Yun M, Park CG, Kim JY, Park HW (2000). "Structural analysis of glyceraldehyde 3-phosphate dehydrogenase from Escherichia coli: direct evidence of substrate binding and cofactor-induced conformational changes." Biochemistry 39(35);10702-10. PMID: 10978154
Zhang07: Zhang N, Chen R, Young N, Wishart D, Winter P, Weiner JH, Li L (2007). "Comparison of SDS- and methanol-assisted protein solubilization and digestion methods for Escherichia coli membrane proteome analysis by 2-D LC-MS/MS." Proteomics 7(4);484-93. PMID: 17309111
Zhang09: Zhang J, Sprung R, Pei J, Tan X, Kim S, Zhu H, Liu CF, Grishin NV, Zhao Y (2009). "Lysine acetylation is a highly abundant and evolutionarily conserved modification in Escherichia coli." Mol Cell Proteomics 8(2);215-25. PMID: 18723842
Zhao95: Zhao G, Pease AJ, Bharani N, Winkler ME (1995). "Biochemical characterization of gapB-encoded erythrose 4-phosphate dehydrogenase of Escherichia coli K-12 and its possible role in pyridoxal 5'-phosphate biosynthesis." J Bacteriol 1995;177(10);2804-12. PMID: 7751290
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