Pathway Tools
Intro Tutorial
discounted registration ends Sept 5, 2015 (Sat)
Pathway Tools
Intro Tutorial
discounted registration ends Sept 5, 2015 (Sat)
Pathway Tools
Intro Tutorial
discounted registration ends Sept 5, 2015 (Sat)
Pathway Tools
Intro Tutorial
discounted registration ends Sept 5, 2015 (Sat)
Pathway Tools
Intro Tutorial
discounted registration ends Sept 5, 2015 (Sat)
twitter

Escherichia coli K-12 substr. MG1655 Enzyme: fructose-1,6-bisphosphatase I



Gene: fbp Accession Numbers: EG10283 (EcoCyc), b4232, ECK4227

Synonyms: fdp

Regulation Summary Diagram: ?

Regulation summary diagram for fbp

Subunit composition of fructose-1,6-bisphosphatase I = [Fbp]4
         fructose-1,6-bisphosphatase monomer = Fbp

Summary:
Fructose-1,6-bisphosphatase catalyzes the conversion of fructose-1,6-bisphosphate to fructose-6-phosphate in the gluconeogenesis pathway. The enzyme is required for growth on glycerol, succinate and acetate as the carbon source, but not for growth on hexoses and pentoses [Fraenkel65].

To avoid futile cycling with phosphofructokinases, the activity of fructose-1,6-bisphosphatase must be regulated. Like the mammalian enzymes, E. coli fructose-1,6-bisphosphatase is inhibited by fructose-2,6-bisphosphate; however, this compound is not present in vivo [KelleyLoughnane02]. AMP is a noncompetitive inhibitor of the enzyme. The low Ki of the enzyme for AMP suggests that under physiological conditions the enzyme would be inhibited to a great extent [Babul83]. A mutant enzyme that is insensitive to AMP inhibition has been isolated [Sedivy86]. Phosphoenolpyruvate (PEP) was initially reported to partially inhibit fructose-1,6-bisphosphatase activity at high concentrations, but was able to block inhibition by AMP [Babul83]. A later report showed that PEP at low concentrations (2 mM) activates the enzyme; PEP may thus be the physiological regulator under gluconeogenic growth conditions [Hines06].

Under nondenaturing conditions, the enzyme is present in several aggregated forms in which the tetramer seems to predominate at low enzyme concentrations [Babul83]. A crystal structure of this enzyme has been solved at 1.45 Å resolution [Hines06]. Crystal structures have also been solved with several bound ligands at 2.05 Å resolution [Hines07] and 2.18 Å resolution [Hines07a, Hines07b].

In addition to PEP, citrate is also an activator. Tetramers with bound PEP or citrate are in the active allosteric R state conformation. PEP, citrate, 3-phosphoglycerate, cis-aconitate, isocitrate, oxaloacetate, and α-ketoglutarate are activators at concentrations of up to 5 mM, although PEP is the most potent activator. Like PEP, citrate is an antagonist of AMP inhibition [Hines07a]. Glucose 6-phosphate is also an allosteric inhibitor and distinct sites for inhibition by AMP and glucose 6-phosphate have been identified. The enzyme undergoes a quaternary structure transition from the R-state to a T-like state in response to the binding of these metabolites. Their binding is proposed to down-regulate the activated R state enzyme [Hines07]. Unlike the porcine enzyme which is synergistically inhibited by AMP and fructose 2,6-bisphosphate, these compounds are not synergistic inhibitors of this Type I E. coli enzyme [Hines07b, Gao14].

Using a kinetic model of E. coli glycolysis, allosteric interactions were identified that govern the reversible switch between glycolysis and gluconeogenesis which included the activation of fructose-1,6-bisphosphatase by pyruvate [Link13]. In E. coli metabolic engineering studies, site-directed mutagenesis of target amino acid residues encoded by fbp reduced allosteric inhibition without affecting catalytic efficiency [Yang12a]. E. coli fbp overexpressed in a strain of Corynebacterium glutamicum resulted in increased L-lysine production [Xu14].

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 fbp at 77% efficacy resulted in a defect in carbon catabolite repression [Nakashima14].

Locations: cytosol

Map Position: [4,452,634 <- 4,453,632] (95.97 centisomes, 345°)
Length: 999 bp / 332 aa

Molecular Weight of Polypeptide: 36.834 kD (from nucleotide sequence), 40 kD (experimental) [Babul83 ]

Molecular Weight of Multimer: 150 kD (experimental) [KelleyLoughnane02]

pI: 5.97

Unification Links: ASAP:ABE-0013842 , CGSC:784 , EchoBASE:EB0279 , EcoGene:EG10283 , EcoliWiki:b4232 , ModBase:P0A993 , OU-Microarray:b4232 , PortEco:fbp , PR:PRO_000022576 , Pride:P0A993 , Protein Model Portal:P0A993 , RefSeq:NP_418653 , RegulonDB:EG10283 , SMR:P0A993 , String:511145.b4232 , Swiss-Model:P0A993 , UniProt:P0A993

Relationship Links: InterPro:IN-FAMILY:IPR000146 , InterPro:IN-FAMILY:IPR020548 , InterPro:IN-FAMILY:IPR028343 , Panther:IN-FAMILY:PTHR11556 , PDB:Structure:2GQ1 , PDB:Structure:2OWZ , PDB:Structure:2OX3 , PDB:Structure:2Q8M , PDB:Structure:2QVR , Pfam:IN-FAMILY:PF00316 , Prints:IN-FAMILY:PR00115 , Prosite:IN-FAMILY:PS00124

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

GO Terms:

Biological Process: GO:0006094 - gluconeogenesis Inferred from experiment Inferred by computational analysis [UniProtGOA12, Fraenkel65]
GO:0016311 - dephosphorylation Inferred from experiment Inferred by computational analysis [GOA06, GOA01, GOA01a, KelleyLoughnane02]
GO:0051289 - protein homotetramerization Inferred from experiment [KelleyLoughnane02]
GO:0005975 - carbohydrate metabolic process Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0008152 - metabolic process Inferred by computational analysis [UniProtGOA11a]
GO:0016051 - carbohydrate biosynthetic process Inferred by computational analysis [GOA06]
Molecular Function: GO:0042132 - fructose 1,6-bisphosphate 1-phosphatase activity Inferred from experiment Inferred by computational analysis [GOA06, GOA01, GOA01a, KelleyLoughnane02]
GO:0000166 - nucleotide binding Inferred by computational analysis [UniProtGOA11a]
GO:0000287 - magnesium ion binding Inferred by computational analysis [GOA06]
GO:0003824 - catalytic activity Inferred by computational analysis [UniProtGOA11a]
GO:0016787 - hydrolase activity Inferred by computational analysis [UniProtGOA11a]
GO:0042578 - phosphoric ester hydrolase activity Inferred by computational analysis [GOA01a]
GO:0046872 - metal ion binding Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Ishihama08, LopezCampistrou05]
GO:0005737 - cytoplasm Inferred by computational analysis [UniProtGOA11, UniProtGOA11a, GOA06]

MultiFun Terms: metabolism central intermediary metabolism

Essentiality data for fbp knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 1]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 2]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 3]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 2]
Yes [Feist07, Comment 4]

Credits:
Curated 20-Dec-2006 by Keseler I , SRI International
Last-Curated ? 19-May-2015 by Fulcher C , SRI International


Enzymatic reaction of: fructose-1,6-bisphosphatase

Synonyms: fructose bisphosphatase, hexosediphosphatase, fructose-1,6-P2 1-phosphatase

EC Number: 3.1.3.11

fructose 1,6-bisphosphate + H2O <=> β-D-fructofuranose 6-phosphate + phosphate

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.

The reaction is physiologically favored in the direction shown.

Alternative Substrates for fructose 1,6-bisphosphate: α-glucose 1,6-bisphosphate [Babul83 ] , β-D-fructofuranose 1-phosphate [Babul83 ]

In Pathways: superpathway of hexitol degradation (bacteria) , 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)

Summary:
The enzyme shows substrate inhibition by fructose1,6-bisphosphate at concentrations above 0.05mM [Babul83].

Values for kinetic parameters were shown to vary with assay protocol [Hines07a].

Cofactors or Prosthetic Groups: Mg2+ [KelleyLoughnane02, Babul83]

Activators (Unknown Mechanism): citrate [Hines07a] , sulfate [Hines06] , phosphoenolpyruvate [Hines06]

Inhibitors (Competitive): β-D-fructose 2,6-bisphosphate [KelleyLoughnane02, Marcus84]

Inhibitors (Noncompetitive): AMP [Babul83]

Inhibitors (Unknown Mechanism): β-D-glucose 6-phosphate [Hines07] , phosphoenolpyruvate [Babul83]

Primary Physiological Regulators of Enzyme Activity: citrate , β-D-glucose 6-phosphate , phosphoenolpyruvate , AMP

Kinetic Parameters:

Substrate
Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)
Citations
fructose 1,6-bisphosphate
5.0
[Babul83]
fructose 1,6-bisphosphate
15.4
14.6
0.95
[KelleyLoughnane02]

pH(opt): 7.5 [Babul83]


Sequence Features

Protein sequence of fructose-1,6-bisphosphatase monomer with features indicated

Feature Class Location Attached Group Citations Comment
Protein-Segment 3 -> 5  
[UniProt10]
UniProt: Allosteric activator binding; Sequence Annotation Type: region of interest;
Nucleotide-Phosphate-Binding-Region 19 -> 23 AMP
[Hines07, UniProt15]
UniProt: AMP.
Amino-Acid-Sites-That-Bind 30  
[UniProt15]
UniProt: Allosteric activator.
Metal-Binding-Site 89  
[UniProt15]
UniProt: Magnesium 1.
Nucleotide-Phosphate-Binding-Region 104 -> 105 AMP
[Hines07, UniProt15]
UniProt: AMP.
Metal-Binding-Site 110  
[UniProt15]
UniProt: Magnesium 1.
Metal-Binding-Site 112  
[UniProt15]
UniProt: Magnesium 1; via carbonyl oxygen.
Metal-Binding-Site 113  
[UniProt15]
UniProt: Magnesium 2.
Protein-Segment 113 -> 116  
[UniProt10]
UniProt: Substrate binding; Sequence Annotation Type: region of interest;
Amino-Acid-Sites-That-Bind 187  
[UniProt15]
UniProt: Allosteric activator; via amide nitrogen.
Amino-Acid-Sites-That-Bind 206  
[UniProt15]
UniProt: Substrate.
Amino-Acid-Sites-That-Bind 222  
[UniProt15]
UniProt: Allosteric inhibitor glucose-6-phosphate.
Amino-Acid-Sites-That-Bind 225  
[UniProt15]
UniProt: Allosteric inhibitor glucose-6-phosphate.
Amino-Acid-Sites-That-Bind 239  
[UniProt15]
UniProt: Substrate.
Protein-Segment 257 -> 259  
[UniProt10]
UniProt: Substrate binding; Sequence Annotation Type: region of interest;
Amino-Acid-Sites-That-Bind 269  
[UniProt15]
UniProt: Substrate.
Metal-Binding-Site 275  
[UniProt15]
UniProt: Magnesium 2.


Gene Local Context (not to scale): ?

Gene local context diagram

Transcription Unit:

Transcription-unit diagram

Notes:

History:
10/20/97 Gene b4232 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10283; confirmed by SwissProt match.


References

Baba06: Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H (2006). "Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection." Mol Syst Biol 2;2006.0008. PMID: 16738554

Babul83: Babul J, Guixe V (1983). "Fructose bisphosphatase from Escherichia coli. Purification and characterization." Arch Biochem Biophys 1983;225(2);944-9. PMID: 6312898

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

Feist07: Feist AM, Henry CS, Reed JL, Krummenacker M, Joyce AR, Karp PD, Broadbelt LJ, Hatzimanikatis V, Palsson BO (2007). "A genome-scale metabolic reconstruction for Escherichia coli K-12 MG1655 that accounts for 1260 ORFs and thermodynamic information." Mol Syst Biol 3;121. PMID: 17593909

Fraenkel65: Fraenkel DG, Horecker BL (1965). "Fructose-1, 6-diphosphatase and acid hexose phosphatase of Escherichia coli." J Bacteriol 90(4);837-42. PMID: 4284917

Gao14: Gao Y, Shen L, Honzatko RB (2014). "Central cavity of fructose-1,6-bisphosphatase and the evolution of AMP/fructose 2,6-bisphosphate synergism in eukaryotic organisms." J Biol Chem 289(12);8450-61. PMID: 24436333

Gerdes03: Gerdes SY, Scholle MD, Campbell JW, Balazsi G, Ravasz E, Daugherty MD, Somera AL, Kyrpides NC, Anderson I, Gelfand MS, Bhattacharya A, Kapatral V, D'Souza M, Baev MV, Grechkin Y, Mseeh F, Fonstein MY, Overbeek R, Barabasi AL, Oltvai ZN, Osterman AL (2003). "Experimental determination and system level analysis of essential genes in Escherichia coli MG1655." J Bacteriol 185(19);5673-84. PMID: 13129938

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."

Hines06: Hines JK, Fromm HJ, Honzatko RB (2006). "Novel allosteric activation site in Escherichia coli fructose-1,6-bisphosphatase." J Biol Chem 281(27);18386-93. PMID: 16670087

Hines07: Hines JK, Kruesel CE, Fromm HJ, Honzatko RB (2007). "Structure of inhibited fructose-1,6-bisphosphatase from Escherichia coli: distinct allosteric inhibition sites for AMP and glucose 6-phosphate and the characterization of a gluconeogenic switch." J Biol Chem 282(34);24697-706. PMID: 17567577

Hines07a: Hines JK, Fromm HJ, Honzatko RB (2007). "Structures of activated fructose-1,6-bisphosphatase from Escherichia coli. Coordinate regulation of bacterial metabolism and the conservation of the R-state." J Biol Chem 282(16);11696-704. PMID: 17314096

Hines07b: Hines JK, Chen X, Nix JC, Fromm HJ, Honzatko RB (2007). "Structures of mammalian and bacterial fructose-1,6-bisphosphatase reveal the basis for synergism in AMP/fructose 2,6-bisphosphate inhibition." J Biol Chem 282(49);36121-31. PMID: 17933867

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

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

KelleyLoughnane02: Kelley-Loughnane N, Biolsi SA, Gibson KM, Lu G, Hehir MJ, Phelan P, Kantrowitz ER (2002). "Purification, kinetic studies, and homology model of Escherichia coli fructose-1,6-bisphosphatase." Biochim Biophys Acta 1594(1);6-16. PMID: 11825604

Link13: Link H, Kochanowski K, Sauer U (2013). "Systematic identification of allosteric protein-metabolite interactions that control enzyme activity in vivo." Nat Biotechnol 31(4);357-61. PMID: 23455438

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

Marcus84: Marcus F, Edelstein I, Rittenhouse J (1984). "Inhibition of Escherichia coli fructose-1,6-bisphosphatase by fructose 2,6-bisphosphate." Biochem Biophys Res Commun 119(3);1103-8. PMID: 6324777

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

Sedivy86: Sedivy JM, Babul J, Fraenkel DG (1986). "AMP-insensitive fructose bisphosphatase in Escherichia coli and its consequences." Proc Natl Acad Sci U S A 83(6);1656-9. PMID: 3006063

UniProt10: UniProt Consortium (2010). "UniProt version 2010-07 released on 2010-06-15 00:00:00." Database.

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

UniProtGOA11: UniProt-GOA (2011). "Gene Ontology annotation based on the manual assignment of UniProtKB Subcellular Location terms in UniProtKB/Swiss-Prot entries."

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

UniProtGOA12: UniProt-GOA (2012). "Gene Ontology annotation based on UniPathway vocabulary mapping."

Xu14: Xu JZ, Zhang JL, Guo YF, Jia QD, Zhang WG (2014). "Heterologous expression of Escherichia coli fructose-1,6-bisphosphatase in Corynebacterium glutamicum and evaluating the effect on cell growth and L-lysine production." Prep Biochem Biotechnol 44(5);493-509. PMID: 24397720

Yang12a: Yang JS, Seo SW, Jang S, Jung GY, Kim S (2012). "Rational engineering of enzyme allosteric regulation through sequence evolution analysis." PLoS Comput Biol 8(7);e1002612. PMID: 22807670


Report Errors or Provide Feedback
Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
Page generated by SRI International Pathway Tools version 19.0 on Wed Sep 2, 2015, biocyc13.