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Catalytic Site Atlas

CSA LITERATURE entry for 2esd

E.C. nameglyceraldehyde-3-phosphate dehydrogenase (NADP+)
SpeciesStreptococcus mutans ()
E.C. Number (IntEnz) 1.2.1.9
CSA Homologues of 2esd
CSA Entries With UniProtID Q59931
CSA Entries With EC Number 1.2.1.9
PDBe Entry 2esd
PDBSum Entry 2esd
MACiE Entry 2esd

Literature Report

IntroductionNADP-dependent glyceraldehyde-3-phosphate dehydrogenase (GAPN) from Streptococcus mutans (GAPN) is a non-phosphorylating, CoA-independent member of the aldehyde dehydrogenase (ALDH) family. It catalyses the irreversible oxidation of glyceraldehyde-3-phosphate (G3P) into 3-phosphoglycerate (3-PGA) in the presence of NADP via a two step mechanism.
MechansimGAPN uses an ordered sequential two-step mechanism for catalysis - acylation followed by deacylation.
First, NADP+ binds to the enzyme, inducing a conformational change in the active site, resulting in the activation of a competent enzyme. The positive charge of the nicotinamide ring of NADP+, the amide peptide nitrogens of residues Cys284 and Thr285, and possibly the involvement of other structural elements stabilise the thiolate form of Cys284.
G3P binding leads to formation of a thiohemiacetal intermediate via a nucleophilic attack by the Cys284 thiolate. This is followed by an oxidoreduction process in which a hydride is transferred from the intermediate to the nicotinamide ring of NADP+ forming a thioacylenzyme intermediate. This is able to take place without catalytic assistance due to the presence of an oxianion hole, formed by Asn154 and the amide nitrogen of Cys284, interacting with the C1 oxygen of the substrate, keeping the oxygen deprotonated. The oxyanion hole is also involved in the stabilisation of the tetrahedral thioacyl intermediate.
Then deacylation occurs, where Glu250 orientates and activates water which nucleophilically attacks the C1 carbon of the thioacyl intermediate, leading to an acid product and, finally, the release of the reduced cofactor (NADPH). The isomerisation and movement of the nicotinamide ring of NADPH after the oxidoreduction stage is required to allow Glu250 to perform its role in deacylation.
Reaction

Catalytic Sites for 2esd

Annotated By Reference To The Literature - Site 1 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
AsnA154154macie:sideChainAsn154-ND2 group and the amide nitrogen of Cys284 interact with the C1 oxygen of the substrate to form an oxianion hole which is required of hydride transfer and involved in the stabilisation of the intermediates.
AlaA250250macie:sideChainGlu250 orientates and activates water which acts as a nucleophile and attacks the C1 carbon of the thioacylenzyme intermediate, leading to the formation an acid product of 3-PGA and the release of the reduced cofactor. At basic pH, deacylation is catalysed by the chemical activation of the water molecule attacking the thioacyl intermediate via the abstraction of a proton by Glu250 acting as a base catalyst.
CysA284284macie:sideChainIn thiolate form, Cys284 acts as a nucleophile and attacks C1 of G3P substrate to form a thiohemiacetal enzyme intermediate in the acylation process. The amide nitrogen of Cys284, in conjunction with Asn154, interacts with the C1 oxygen of the substrate to form an oxianion hole which is required for hydride transfer and involved in the stabilisation of the intermediates.
CysA284284macie:mainChainAmideIn thiolate form, Cys284 acts as a nucleophile and attacks C1 of G3P substrate to form a thiohemiacetal enzyme intermediate in the acylation process. The amide nitrogen of Cys284, in conjunction with Asn154, interacts with the C1 oxygen of the substrate to form an oxianion hole which is required for hydride transfer and involved in the stabilisation of the intermediates.

Annotated By Reference To The Literature - Site 2 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
AsnB154154macie:sideChainAsn154-ND2 group and the amide nitrogen of Cys284 interact with the C1 oxygen of the substrate to form an oxianion hole which is required of hydride transfer and involved in the stabilisation of the intermediates.
CysB284284macie:sideChainIn thiolate form, Cys284 acts as a nucleophile and attacks C1 of G3P substrate to form a thiohemiacetal enzyme intermediate in the acylation process. The amide nitrogen of Cys284, in conjunction with Asn154, interacts with the C1 oxygen of the substrate to form an oxianion hole which is required for hydride transfer and involved in the stabilisation of the intermediates.
CysB284284macie:mainChainAmideIn thiolate form, Cys284 acts as a nucleophile and attacks C1 of G3P substrate to form a thiohemiacetal enzyme intermediate in the acylation process. The amide nitrogen of Cys284, in conjunction with Asn154, interacts with the C1 oxygen of the substrate to form an oxianion hole which is required for hydride transfer and involved in the stabilisation of the intermediates.
AlaB250250macie:sideChainGlu250 orientates and activates water which acts as a nucleophile and attacks the C1 carbon of the thioacylenzyme intermediate, leading to the formation an acid product of 3-PGA and the release of the reduced cofactor. At basic pH, deacylation is catalysed by the chemical activation of the water molecule attacking the thioacyl intermediate via the abstraction of a proton by Glu250 acting as a base catalyst.

Annotated By Reference To The Literature - Site 3 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
AsnC154154macie:sideChainAsn154-ND2 group and the amide nitrogen of Cys284 interact with the C1 oxygen of the substrate to form an oxianion hole which is required of hydride transfer and involved in the stabilisation of the intermediates.
CysC284284macie:sideChainIn thiolate form, Cys284 acts as a nucleophile and attacks C1 of G3P substrate to form a thiohemiacetal enzyme intermediate in the acylation process. The amide nitrogen of Cys284, in conjunction with Asn154, interacts with the C1 oxygen of the substrate to form an oxianion hole which is required for hydride transfer and involved in the stabilisation of the intermediates.
CysC284284macie:mainChainAmideIn thiolate form, Cys284 acts as a nucleophile and attacks C1 of G3P substrate to form a thiohemiacetal enzyme intermediate in the acylation process. The amide nitrogen of Cys284, in conjunction with Asn154, interacts with the C1 oxygen of the substrate to form an oxianion hole which is required for hydride transfer and involved in the stabilisation of the intermediates.
AlaC250250macie:sideChainGlu250 orientates and activates water which acts as a nucleophile and attacks the C1 carbon of the thioacylenzyme intermediate, leading to the formation an acid product of 3-PGA and the release of the reduced cofactor. At basic pH, deacylation is catalysed by the chemical activation of the water molecule attacking the thioacyl intermediate via the abstraction of a proton by Glu250 acting as a base catalyst.

Annotated By Reference To The Literature - Site 4 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
AsnD154154macie:sideChainAsn154-ND2 group and the amide nitrogen of Cys284 interact with the C1 oxygen of the substrate to form an oxianion hole which is required of hydride transfer and involved in the stabilisation of the intermediates.
CysD284284macie:sideChainIn thiolate form, Cys284 acts as a nucleophile and attacks C1 of G3P substrate to form a thiohemiacetal enzyme intermediate in the acylation process. The amide nitrogen of Cys284, in conjunction with Asn154, interacts with the C1 oxygen of the substrate to form an oxianion hole which is required for hydride transfer and involved in the stabilisation of the intermediates.
CysD284284macie:mainChainAmideIn thiolate form, Cys284 acts as a nucleophile and attacks C1 of G3P substrate to form a thiohemiacetal enzyme intermediate in the acylation process. The amide nitrogen of Cys284, in conjunction with Asn154, interacts with the C1 oxygen of the substrate to form an oxianion hole which is required for hydride transfer and involved in the stabilisation of the intermediates.
AlaD250250macie:sideChainGlu250 orientates and activates water which acts as a nucleophile and attacks the C1 carbon of the thioacylenzyme intermediate, leading to the formation an acid product of 3-PGA and the release of the reduced cofactor. At basic pH, deacylation is catalysed by the chemical activation of the water molecule attacking the thioacyl intermediate via the abstraction of a proton by Glu250 acting as a base catalyst.

Literature References

Notes:
D'Ambrosio K
The first crystal structure of a thioacylenzyme intermediate in the ALDH family: new coenzyme conformation and relevance to catalysis.
Biochemistry 2006 45 2978-2986
PubMed: 16503652
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