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Catalytic Site Atlas

CSA LITERATURE entry for 1db3

E.C. nameGDP-mannose 4,6-dehydratase
SpeciesEscherichia coli (Bacteria)
E.C. Number (IntEnz)
CSA Homologues of 1db3There are 65 Homologs
CSA Entries With UniProtID P0AC88
CSA Entries With EC Number
PDBe Entry 1db3
PDBSum Entry 1db3
MACiE Entry 1db3

Literature Report

IntroductionGDP-mannose 4,6-dehydratase (GMD) catalyses the conversion of GDP-mannose to GDP-4-dehydro-6deoxy-D-mannose in the first and regulatory step in the de novo biosynthesis of GDP-fucose. Fucose is distributed widely in the glycoconjugates of species ranging from bacteria to humans. Once synthesised it is incorporated into glycoconjugates by specific fucosyltransferases that use GDP-fucose as the fucose donor. Important glycoconjugates include the cell wall polysaccharides of bacteria that confer pathogenicity, components of mammal antigens and perhaps the best known example, the human blood group antigens. Defects in GMD activity provide insight into the diverse roles that fucose plays in biology and hence the importance of the enzyme. For example in plants, defects in GMD result in altered shoot development whilst in humans defects have been linked to immune disorders.
MechansimFirstly a basic group on the enzyme, Tyr156, abstracts a proton from the C4 hydroxyl group followed by hydride transfer to the nicotinamide ring of NADP resulting in oxidation/dehydrogenationn of the substrate and formation of NADPH. Then a proton is abstracted from the C5 position by a second base, Glu134, followed by syn elimination of a water molecule between C5 and C6 to give a keto-5,6 manoseen species. The final step involves a hydride transfer from NADPH to yield GDP-4-dehydro-6-deoxy-D-mannose and regeneration of NADP. Thr132 is thought to act as a proton shuffle to Tyr156. Lys160 is thought to coordinate the ribose hydroxyls of NADP as well as lowering the pKa of Tyr156, helping it to abstract protons.
A second mechanism has been proposed, involving the formation of a 4-enediol/enolate species as an intermediate in the second step. The first step is as above. Step two involves the enolisation of the 4-keto group and abstraction of the C5 of the sugar by the second active site base, Glu134. Finally the hydride is transferred from NADPH to the C6 carbon followed by elimination of the water molecule at the C6 position and reprotonation of C5 by Glu134. Thr132 is thought to act as a proton shuffle in both mechanisms. Lys160 coordinates the ribose hydroxyls of NADP as well as lowering the pKa of Tyr156, helping it to abstract protons.
Kinetic analysis, however, gives evidence to suggest that the role of NADP+ does not support either of the above mechanisms. The exact overall mechanism remains unclear.

Catalytic Sites for 1db3

Annotated By Reference To The Literature - Site 1 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
GluA134135macie:sideChainThe residue acts as a general base towards the mannose C5 atom during dehydration and then as a general acid to the 5,6 mannoseen intermediate during hydride transfer to C6 from NADPH.
LysA160161macie:sideChainThe electrostatic interaction between Lys150 and Tyr156 lowers the phenolic oxygen pKa, allowing the anionic form to exist in pH conditions suited to the active site. It is also thought to coordinate to the ribose hydroxyls of NADP.
ThrA132133macie:sideChainThe residue is thought to participate in a proton relay with the catalytic base Tyr156.
TyrA156157macie:sideChain The phenolic oxygen is stabilised in its anionic form by the electrostatic interaction created between the oxygen and Lys160. This activates Tyr156 to act as a base towards the C1-OH group of the mannose substrate. The residue is thought to participate in a proton relay with Thr132.

Literature References

Nasarre P
Guidance molecules in lung cancer.
Cell Adh Migr 2010 4 130-145
PubMed: 20139699
King JD
The structural basis for catalytic function of GMD and RMD, two closely related enzymes from the GDP-D-rhamnose biosynthesis pathway.
FEBS J 2009 276 2686-2700
PubMed: 19459932
Fruscione F
Differential role of NADP+ and NADPH in the activity and structure of GDP-D-mannose 4,6-dehydratase from two chlorella viruses.
J Biol Chem 2008 283 184-193
PubMed: 17974560