Glyceraldehyde-3-phosphate dehydrogenase (NAD(P)+) (phosphorylating)

 

Archaebacterial glyceraldehyde-3-phosphate dehydrogenase is able to catalyse the conversion of glycerate-1-3-bisphophate to glyceraldehyde-3-phosphate using NADPH or NADH as the cofactor. It displays homology with the family of GAPDH dehydrogenases including both bacterial and human forms, but is unique in its resistance to extremes of temperature.

 

Reference Protein and Structure

Sequence
P10618 UniProt (1.2.1.59) IPR006436 (Sequence Homologues) (PDB Homologues)
Biological species
Methanothermus fervidus (Archaea) Uniprot
PDB
1cf2 - THREE-DIMENSIONAL STRUCTURE OF D-GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE FROM THE HYPERTHERMOPHILIC ARCHAEON METHANOTHERMUS FERVIDUS (2.1 Å) PDBe PDBsum 1cf2
Catalytic CATH Domains
3.30.360.10 CATHdb (see all for 1cf2)
Click To Show Structure

Enzyme Reaction (EC:1.2.1.59)

hydrogenphosphate
CHEBI:43474ChEBI
+
D-glyceraldehyde 3-phosphate(2-)
CHEBI:59776ChEBI
+
NAD(1-)
CHEBI:57540ChEBI
→
3-phosphonato-D-glyceroyl phosphate(4-)
CHEBI:57604ChEBI
+
hydron
CHEBI:15378ChEBI
+
NADH(2-)
CHEBI:57945ChEBI
Alternative enzyme names: Triosephosphate dehydrogenase (NAD(P)), NAD(P)-dependent glyceraldehyde-3-phosphate dehydrogenase, Triosephosphate dehydrogenase (NAD(P)(+)),

Enzyme Mechanism

Introduction

The reaction proceeds via initial nucleophilic addition-elimination by cysteine on glycerate-1-3-bisphosphate resulting in the thioacyl enzyme intermediate. Hydride transfer to the intermediate from the cofactor results in a further hemithiolaceto enzyme intermediate, with protonation by histidine stabilising this state. Collapse of this intermediate results in the product being released.

Catalytic Residues Roles

UniProt PDB* (1cf2)
Cys140 Cys140P(A) Acts as nucleophile to attack substrate resulting in the elimination fo the phosphate group and formation of the thioacyl-enzyme intermediate which then collapses to give the product. covalently attached, nucleofuge, nucleophile, proton acceptor, proton donor
His219 His219P(A) Acts to increase nucleophilicity of cys 140 by accepting a proton. Also acts to allow hydride transfer to the substrate by stabilising the hemithioacetal intermediate through hydrogen bonding. Finally protonates the cysteine residue to facilitate collapse of this intermediate. proton acceptor, modifies pKa, electrostatic stabiliser, proton donor
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

proton transfer, bimolecular nucleophilic addition, overall reactant used, intermediate formation, enzyme-substrate complex formation, aromatic bimolecular nucleophilic addition, hydride transfer, intermediate collapse, cofactor used, unimolecular elimination by the conjugate base, overall product formed, enzyme-substrate complex cleavage, inferred reaction step, native state of enzyme regenerated

References

  1. Charron C et al. (2000), J Mol Biol, 297, 481-500. The crystal structure of d-glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic archaeon Methanothermus fervidus in the presence of NADP+ at 2.1 Å resolution. DOI:10.1006/jmbi.2000.3565. PMID:10715215.

Step 1. His219 decreases the pKa of Cys140 and deprotonates the thiol group.

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Catalytic Residues Roles

Residue Roles
His219P(A) modifies pKa, proton acceptor
Cys140P(A) proton donor

Chemical Components

proton transfer

Step 2. Cys140 attacks D-glyceraldehyde in a nucleophilic bimolecular addition reaction.

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Catalytic Residues Roles

Residue Roles
Cys140P(A) covalently attached, nucleophile

Chemical Components

ingold: bimolecular nucleophilic addition, overall reactant used, intermediate formation, enzyme-substrate complex formation

Step 3. A hydride is transferred from the thioacyl enzyme intermediate to NAD and the tetrahedral intermediate collapses. Hydride transfer is promoted by His219, either as a base catalyst or as a stabilizer of the deprotonated hemithioacetal intermediate.

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Catalytic Residues Roles

Residue Roles
Cys140P(A) covalently attached
His219P(A) electrostatic stabiliser

Chemical Components

ingold: aromatic bimolecular nucleophilic addition, hydride transfer, intermediate collapse, cofactor used

Step 4. The phosphate attacks the intermediate in a nucleophilic addition reaction.

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Catalytic Residues Roles

Residue Roles
Cys140P(A) covalently attached
His219P(A) electrostatic stabiliser

Chemical Components

ingold: bimolecular nucleophilic addition, intermediate formation

Step 5. The tetrahedral intermediate collapses, forming the product and eliminating Cys140.

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Catalytic Residues Roles

Residue Roles
Cys140P(A) nucleofuge

Chemical Components

ingold: unimolecular elimination by the conjugate base, intermediate collapse, overall product formed, enzyme-substrate complex cleavage

Step 6. In an inferred step the native state of the active site is regenerated after proton transfer to Cys140 and from His219.

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Catalytic Residues Roles

Residue Roles
His219P(A) proton donor
Cys140P(A) proton acceptor

Chemical Components

proton transfer, inferred reaction step, native state of enzyme regenerated
Download: Image, Marvin File

Step 1. His219 decreases the pKa of Cys140 and deprotonates the thiol group.

Step 2. Cys140 attacks D-glyceraldehyde in a nucleophilic bimolecular addition reaction.

Step 3. A hydride is transferred from the thioacyl enzyme intermediate to NAD and the tetrahedral intermediate collapses. Hydride transfer is promoted by His219, either as a base catalyst or as a stabilizer of the deprotonated hemithioacetal intermediate.

Step 4. The phosphate attacks the intermediate in a nucleophilic addition reaction.

Step 5. The tetrahedral intermediate collapses, forming the product and eliminating Cys140.

Step 6. In an inferred step the native state of the active site is regenerated after proton transfer to Cys140 and from His219.

Products.

Contributors

Henry Pertinez, Peter Sarkies, Gemma L. Holliday, Amelia Brasnett