Anthocyanidin synthase

 

The 2-oxoglutarate dependent enzyme anthocyanidin synthase is able to catalyse the synthesis of anthocyanidin from quercetin, a vital step in the synthesis of flavoids and flavones in plant cells. The reaction is coupled to the decarboxylation of 2-oxoglutarate to form succinate. It is part of the family of 2OG dependent non-haem iron oxygenases, which use 2-oxoglutarate as a cosubstrate and Iron (II) as a cofactor in their reaction mechanisms. The enzyme is of particular interest to geneticists as a mutated form is responsible for the original tall/dwarf dimorphism observed in peas by Mendel.

 

Reference Protein and Structure

Sequence
Q96323 UniProt (1.14.20.4) IPR027443 (Sequence Homologues) (PDB Homologues)
Biological species
Arabidopsis thaliana (Thale cress) Uniprot
PDB
1gp5 - Anthocyanidin synthase from Arabidopsis thaliana complexed with trans-dihydroquercetin (2.2 Å) PDBe PDBsum 1gp5
Catalytic CATH Domains
2.60.120.330 CATHdb (see all for 1gp5)
Cofactors
Iron(3+) (1)
Click To Show Structure

Enzyme Reaction (EC:1.14.20.4)

2-oxoglutarate(2-)
CHEBI:16810ChEBI
+
(2R,3S,4S)-leucocyanidin
CHEBI:11412ChEBI
+
dioxygen
CHEBI:15379ChEBI
→
water
CHEBI:15377ChEBI
+
succinate(2-)
CHEBI:30031ChEBI
+
carbon dioxide
CHEBI:16526ChEBI
+
(+)-taxifolin
CHEBI:17948ChEBI
Alternative enzyme names: Leucoanthocyanidin dioxygenase, Leucocyanidin oxygenase, Leucocyanidin,2-oxoglutarate:oxygen oxidoreductase, ANS (gene name),

Enzyme Mechanism

Introduction

In the first stage of the reaction the iron (II) ion acts as a nucleophile to attack molecular oxygen, thus reducing it to a peroxide radical, and oxidising iron (II) to iron (III). The radical then reacts with the 2C of the 2 oxoglutarate to create an unstable intermediate which loses CO2 to give iron IV with a double bond formed to oxygen. The reduction of iron to Fe (III) then allows the double bond to break, so that the oxygen can strip a proton from the substrate quercetin to leave a radical. The radical then allows the OH to break its bond to the iron, forming iron (II) and an acetal. Deprotonation of the acetal by Lys 213 results in the return of the OH to the Iron (II), now acting as a Lewis acid, and forms the product Anthocyanidin.

Catalytic Residues Roles

UniProt PDB* (1gp5)
His232, His288, Asp234 His232A, His288A, Asp234A The residues are coordinated to the Fe(II) ion centre. metal ligand
Lys213 Lys213A Deprotonates quercetin to allow a double bond to form so that dehydration occurs resulting in the Anthrocyadin product. proton relay, proton acceptor, 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

electron transfer, coordination to a metal ion, intermediate formation, decarboxylation, overall product formed, intermediate collapse, radical formation, radical termination, bimolecular elimination, keto-enol tautomerisation, native state of enzyme regenerated

References

  1. Wilmouth RC et al. (2002), Structure, 10, 93-103. Structure and Mechanism of Anthocyanidin Synthase from Arabidopsis thaliana. DOI:10.1016/s0969-2126(01)00695-5. PMID:11796114.

Step 1. Fe(II) transfers an electron to the triplet state dioxygen.

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Catalytic Residues Roles

Residue Roles
His232A metal ligand
Asp234A metal ligand
His288A metal ligand

Chemical Components

electron transfer, coordination to a metal ion

Step 2. The superoxide attacks the keto group of 2-oxoglutarate.

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Catalytic Residues Roles

Residue Roles
His232A metal ligand
Asp234A metal ligand
His288A metal ligand

Chemical Components

intermediate formation, electron transfer

Step 3. There is oxidative decarboxylation, resulting in the formation of succinate, CO2 and the ferryl species.

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Catalytic Residues Roles

Residue Roles
His232A metal ligand
Asp234A metal ligand
His288A metal ligand

Chemical Components

decarboxylation, overall product formed, intermediate collapse

Step 4. The ferryl oxygen abstracts a proton from leucocyanidin to form a radical intermediate.

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Catalytic Residues Roles

Residue Roles
His232A metal ligand
Asp234A metal ligand
His288A metal ligand

Chemical Components

radical formation, intermediate formation

Step 5. The hydroxide adds to the radical intermediate, reducing Fe(III) to Fe(II) and forming an acetal.

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Catalytic Residues Roles

Residue Roles
His232A metal ligand
Asp234A metal ligand
His288A metal ligand

Chemical Components

radical termination, intermediate formation

Step 6. Lys213 acts as a base to deprotonate the acetal intermediate, causing the elimination of the hydroxyl and formation of a carbon-carbon double bond.

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Catalytic Residues Roles

Residue Roles
His232A metal ligand
Asp234A metal ligand
His288A metal ligand
Lys213A proton acceptor

Chemical Components

ingold: bimolecular elimination, overall product formed

Step 7. In an inferred step the product of step 6 tautomerises to form the taxifolin product. The succinate product and hydroxyl are released from the Fe(II) centre and the native state of the enzyme is restored.

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Catalytic Residues Roles

Residue Roles
His232A metal ligand
Asp234A metal ligand
His288A metal ligand
Lys213A proton relay, proton donor, proton acceptor

Chemical Components

keto-enol tautomerisation, native state of enzyme regenerated
Download: Image, Marvin File

Step 1. Fe(II) transfers an electron to the triplet state dioxygen.

Step 2. The superoxide attacks the keto group of 2-oxoglutarate.

Step 3. There is oxidative decarboxylation, resulting in the formation of succinate, CO2 and the ferryl species.

Step 4. The ferryl oxygen abstracts a proton from leucocyanidin to form a radical intermediate.

Step 5. The hydroxide adds to the radical intermediate, reducing Fe(III) to Fe(II) and forming an acetal.

Step 6. Lys213 acts as a base to deprotonate the acetal intermediate, causing the elimination of the hydroxyl and formation of a carbon-carbon double bond.

Step 7. In an inferred step the product of step 6 tautomerises to form the taxifolin product. The succinate product and hydroxyl are released from the Fe(II) centre and the native state of the enzyme is restored.

Products.

Contributors

Peter Sarkies, Gemma L. Holliday, Amelia Brasnett