Escherichia coli K-12 substr. MG1655 Enzyme: GDP-mannose 4,6-dehydratase

Gene: gmd Accession Numbers: EG11787 (EcoCyc), b2053, ECK2047

Synonyms: yefN, yefA

Regulation Summary Diagram: ?

Regulation summary diagram for gmd

Subunit composition of GDP-mannose 4,6-dehydratase = [Gmd]2
         GDP-mannose 4,6-dehydratase = Gmd

GDP-mannose 4,6-dehydratase catalyzes the first step in the pathway that converts GDP-D-mannose to GDP-L-fucose. GDP-L-fucose is one of the precursors of colanic acid (M antigen), an extracellular polysaccharide [Sturla97, Stevenson96].

Recombinant enzyme was expressed and purified as a GST fusion protein and characterized both with and without the GST tag. These studies suggested that the active form of the enzyme is a homohexamer, containing one mole of tightly bound NADP per monomer [Sturla97]. However, later work showed the enzyme to be a homodimer [Somoza00].

The crystal structure of the enzyme has been determined at 2.3 Å resolution. It is a member of the short-chain dehydrogenase/reductase (SDR) family. Each monomer was shown to be composed of a larger N-terminal domain that binds the NADP or NADPH cofactor, and a C-terminal domain with the sugar-nucleotide binding site [Somoza00].

This enzyme as also been used in metabolic engineering studies to produce GDP-L-fucose for the synthesis of fucosylated oligosaccharides [Albermann01, Byun07].

Review: [Tonetti98a]

Locations: cytosol

Map Position: [2,125,217 <- 2,126,338] (45.81 centisomes, 165°)
Length: 1122 bp / 373 aa

Molecular Weight of Polypeptide: 42.047 kD (from nucleotide sequence)

Unification Links: ASAP:ABE-0006803 , DIP:DIP-48216N , EchoBASE:EB1735 , EcoGene:EG11787 , EcoliWiki:b2053 , ModBase:P0AC88 , OU-Microarray:b2053 , PortEco:gmd , PR:PRO_000022819 , Pride:P0AC88 , Protein Model Portal:P0AC88 , RefSeq:NP_416557 , RegulonDB:EG11787 , SMR:P0AC88 , String:511145.b2053 , UniProt:P0AC88

Relationship Links: InterPro:IN-FAMILY:IPR001509 , InterPro:IN-FAMILY:IPR006368 , InterPro:IN-FAMILY:IPR016040 , PDB:Structure:1DB3 , Pfam:IN-FAMILY:PF01370

In Paralogous Gene Group: 191 (7 members)

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

GO Terms:

Biological Process: GO:0009242 - colanic acid biosynthetic process Inferred by computational analysis [UniProtGOA12]
GO:0019673 - GDP-mannose metabolic process Inferred by computational analysis [GOA01a]
GO:0042351 - 'de novo' GDP-L-fucose biosynthetic process Inferred by computational analysis [UniProtGOA12]
Molecular Function: GO:0008446 - GDP-mannose 4,6-dehydratase activity Inferred from experiment Inferred by computational analysis [GOA06, GOA01, GOA01a, Sturla97]
GO:0042803 - protein homodimerization activity Inferred from experiment [Somoza00]
GO:0003824 - catalytic activity Inferred by computational analysis [GOA01a]
GO:0016829 - lyase activity Inferred by computational analysis [UniProtGOA11a]
GO:0050662 - coenzyme binding Inferred by computational analysis [GOA01a]
GO:0070401 - NADP+ binding Inferred by computational analysis [GOA06]
Cellular Component: GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

MultiFun Terms: cell processes adaptations dessication
cell structure capsule (M and K antigens)
cell structure surface antigens (ECA, O antigen of LPS)
metabolism biosynthesis of building blocks cofactors, small molecule carriers nicotinamide adenine dinucleotide
metabolism biosynthesis of macromolecules (cellular constituents) colanic acid (M antigen)

Essentiality data for gmd 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]

Last-Curated ? 24-Jan-2013 by Fulcher C , SRI International

Enzymatic reaction of: GDP-mannose 4,6-dehydratase

Synonyms: GDP-D-mannose 4-oxido-6-reductase, GDP-mannose 4,6-hydro-lyase

EC Number:

GDP-α-D-mannose <=> GDP-4-dehydro-6-deoxy-α-D-mannose + H2O

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction of enzyme catalysis.

The reaction is irreversible in the direction shown. [Elbein65]

In Pathways: colanic acid building blocks biosynthesis , GDP-L-fucose biosynthesis I (from GDP-D-mannose)

The enzyme from E. coli 0111-B4 was inhibited by p-chloromercuriphenylsulfonate. The GDP-4-dehydro-6-deoxy-D-mannose product of the reaction is unstable [Elbein65].

Cofactors or Prosthetic Groups: NADP+ [Comment 5, Sturla97]

Activators (Unknown Mechanism): Ca2+ [Sturla97] , Mg2+ [Sturla97]

Inhibitors (Competitive): GDP-L-fucose [Somoza00, Comment 6]

Inhibitors (Unknown Mechanism): GDP-α-D-glucose [Sturla97, Comment 7] , p-chloromercuriphenylsulfonate [Elbein65]

Primary Physiological Regulators of Enzyme Activity: GDP-α-D-glucose , GDP-L-fucose

Kinetic Parameters:

Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)
[Sullivan98, BRENDA14]
[Elbein65, BRENDA14]
222.0, 220.0
[Sturla97, BRENDA14]
[Somoza00, BRENDA14]

pH(opt) (forward direction): 8.0 [BRENDA14, Sturla97]

pH(opt): 7 [BRENDA14, Elbein65]

Sequence Features

Protein sequence of GDP-mannose 4,6-dehydratase with features indicated

Feature Class Location Attached Group Citations Comment
Nucleotide-Phosphate-Binding-Region 9 -> 14 NADP+
UniProt: NADP; Non-Experimental Qualifier: by similarity.
Nucleotide-Phosphate-Binding-Region 64 -> 65 NADP+
UniProt: NADP; Non-Experimental Qualifier: by similarity.
Nucleotide-Phosphate-Binding-Region 86 -> 90 NADP+
UniProt: NADP; Non-Experimental Qualifier: by similarity.
Amino-Acid-Sites-That-Bind 101  
UniProt: NADP; Non-Experimental Qualifier: by similarity.
Active-Site 133  
Mutagenesis-Variant 133  
[Somoza00, UniProt11]
UniProt: Strongly reduces activity.
Mutagenesis-Variant 135  
UniProt: Strongly reduces activity.
Active-Site 135  
UniProt: Nucleophile.
Active-Site 157  
UniProt: Nucleophile.
Mutagenesis-Variant 157  
[Somoza00, UniProt11]
UniProt: Strongly reduces activity.
Amino-Acid-Sites-That-Bind 161  
UniProt: NADP; Non-Experimental Qualifier: probable.
Mutagenesis-Variant 161  
[Somoza00, UniProt11]
UniProt: Strongly reduces activity.
Amino-Acid-Sites-That-Bind 187  
UniProt: NADP; Non-Experimental Qualifier: by similarity.
Amino-Acid-Sites-That-Bind 192  
UniProt: NADP; Non-Experimental Qualifier: by similarity.

Gene Local Context (not to scale): ?

Gene local context diagram

Transcription Unit:

Transcription-unit diagram


10/20/97 Gene b2053 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG11787; confirmed by SwissProt match.


Albermann01: Albermann C, Piepersberg W, Wehmeier UF (2001). "Synthesis of the milk oligosaccharide 2'-fucosyllactose using recombinant bacterial enzymes." Carbohydr Res 334(2);97-103. PMID: 11502265

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

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

Byun07: Byun SG, Kim MD, Lee WH, Lee KJ, Han NS, Seo JH (2007). "Production of GDP-L-fucose, L-fucose donor for fucosyloligosaccharide synthesis, in recombinant Escherichia coli." Appl Microbiol Biotechnol 74(4);768-75. PMID: 17111133

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

Elbein65: Elbein AD, Heath EC (1965). "The Biosynthesis of Cell Wall Lipopolysaccharide in Escherichia coli. II. Guanosine diphosphate 4-keto-6-deoxy-D-mannose, an intermediate in the biosynthesis of guanosine diphosphate colitose." J Biol Chem 240:1926-1931. PMID: 14299611

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

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."

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

Somoza00: Somoza JR, Menon S, Schmidt H, Joseph-McCarthy D, Dessen A, Stahl ML, Somers WS, Sullivan FX (2000). "Structural and kinetic analysis of Escherichia coli GDP-mannose 4,6 dehydratase provides insights into the enzyme's catalytic mechanism and regulation by GDP-fucose." Structure Fold Des 2000;8(2);123-35. PMID: 10673432

Stevenson96: Stevenson G, Andrianopoulos K, Hobbs M, Reeves PR (1996). "Organization of the Escherichia coli K-12 gene cluster responsible for production of the extracellular polysaccharide colanic acid." J Bacteriol 1996;178(16);4885-93. PMID: 8759852

Sturla97: Sturla L, Bisso A, Zanardi D, Benatti U, De Flora A, Tonetti M (1997). "Expression, purification and characterization of GDP-D-mannose 4,6-dehydratase from Escherichia coli." FEBS Lett 1997;412(1);126-30. PMID: 9257704

Sullivan98: Sullivan FX, Kumar R, Kriz R, Stahl M, Xu GY, Rouse J, Chang XJ, Boodhoo A, Potvin B, Cumming DA (1998). "Molecular cloning of human GDP-mannose 4,6-dehydratase and reconstitution of GDP-fucose biosynthesis in vitro." J Biol Chem 273(14);8193-202. PMID: 9525924

Tonetti98a: Tonetti M, Sturla L, Bisso A, Zanardi D, Benatti U, De Flora A (1998). "The metabolism of 6-deoxyhexoses in bacterial and animal cells." Biochimie 80(11);923-31. PMID: 9893952

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

UniProt13: UniProt Consortium (2013). "UniProt version 2013-08 released on 2013-08-01 00:00:00." Database.

UniProt14: UniProt Consortium (2014). "UniProt version 2014-01 released on 2014-01-01 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."

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

Webb92: Webb EC "Enzyme Nomenclature 1992." Academic Press, Inc., San Diego, 1992.

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Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
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