2-hydroxychromene-2-carboxylate isomerase

 

Catalyzes the reversible glutathione-dependent isomerization of 2-hydroxychromene-2-carboxylate (HCCA) to trans-O-hydroxybenzylidenepyruvate (THBPA). HCCA isomerases are present in most aerobic organisms including microorganisms, plants, and animals. The group kappa GSH transferases (k-GSTs) are a close relative of the largest group of proteins in nature; the GST group. k-GSTs are dimers with a main catalytic domain, which is the N-terminal domain containing the thioredoxin domain, and the C-terminal domain which is mostly alfa-helical and unlike its GST relatives, inserted into the N-terminal domain. HCCA is a part of the naphthalene catabolic pathway of Pseudomonas putida.

 

Reference Protein and Structure

Sequence
Q51948 UniProt (5.99.1.4) IPR044087 (Sequence Homologues) (PDB Homologues)
Biological species
Pseudomonas putida (Bacteria) Uniprot
PDB
2ime - 2-Hydroxychromene-2-carboxylate Isomerase: a Kappa Class Glutathione-S-Transferase from Pseudomonas putida (1.7 Å) PDBe PDBsum 2ime
Catalytic CATH Domains
3.40.30.10 CATHdb (see all for 2ime)
Cofactors
Glutathione (1)
Click To Show Structure

Enzyme Reaction (EC:5.99.1.4)

2-hydroxychromene-2-carboxylate
CHEBI:59350ChEBI
→
(3E)-4-(2-hydroxyphenyl)-2-oxobut-3-enoate
CHEBI:59353ChEBI
Alternative enzyme names: HCCA isomerase, 2HC2CA isomerase, 2-hydroxychromene-2-carboxylic acid isomerase,

Enzyme Mechanism

Introduction

Ser11 deprotonates GSH which then turns into a nucleophile. The nucleophilic sulphur atom attacks C7 of the substrate causing a isomerisation and rearrangement of the molecule. The end of this rearrangement causes the GSH to cleave its S-C connection to the substrate, ending the reaction. GSH is bound to Val168, Trp179, Asp182 and Asn181. The research indicates the stabilisation of GSH by these residues is a major contributor to the reaction.

Catalytic Residues Roles

UniProt PDB* (2ime)
Ser11 Ser11A Activates GSH by removing a proton from its reactive sulphur atom. activator, proton acceptor
Trp179, Asp182, Asn181, Val168 (main) Trp179A, Asp182A, Asn181A, Val168A (main) These residues bind GSH and stabilise the structure so it can react with the substrate. electrostatic stabiliser
*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, michael addition, overall reactant used, intramolecular rearrangement

References

  1. Thompson LC et al. (2007), Biochemistry, 46, 6710-6722. 2-Hydroxychromene-2-carboxylic acid isomerase: a kappa class glutathione transferase from Pseudomonas putida. DOI:10.1021/bi700356u. PMID:17508726.
  2. Deponte M (2013), 1830, 3217-3266. Glutathione catalysis and the reaction mechanisms of glutathione-dependent enzymes. DOI:https://doi.org/10.1016/j.bbagen.2012.09.018.
  3. Ladner JE et al. (2004), Biochemistry, 43, 352-361. Parallel evolutionary pathways for glutathione transferases: structure and mechanism of the mitochondrial class kappa enzyme rGSTK1-1. DOI:10.1021/bi035832z. PMID:14717589.

Step 1. Ser11 deprotonates and activates GSH. GSH is more likey to release a proton due to residues interacting with it. The residues anchoring GSH are essential for this reaction.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Ser11A activator, proton acceptor
Trp179A electrostatic stabiliser
Asp182A electrostatic stabiliser
Asn181A electrostatic stabiliser
Val168A (main) electrostatic stabiliser

Chemical Components

proton transfer

Step 2. GSH bonds its nucleophilic sulphur to the C7 atom of the sustrate.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Trp179A electrostatic stabiliser
Asp182A electrostatic stabiliser
Asn181A electrostatic stabiliser
Val168A (main) electrostatic stabiliser

Chemical Components

michael addition, overall reactant used

Step 3. The proton transfer in the substrate is inferred by the curator. The substrate rotates and is rearranged. This bond rearrangement causes a bond cleavage between the substrate and GSH.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Trp179A electrostatic stabiliser
Asp182A electrostatic stabiliser
Asn181A electrostatic stabiliser
Val168A (main) electrostatic stabiliser

Chemical Components

proton transfer, intramolecular rearrangement
Download: Image, Marvin File

Step 1. Ser11 deprotonates and activates GSH. GSH is more likey to release a proton due to residues interacting with it. The residues anchoring GSH are essential for this reaction.

Step 2. GSH bonds its nucleophilic sulphur to the C7 atom of the sustrate.

Step 3. The proton transfer in the substrate is inferred by the curator. The substrate rotates and is rearranged. This bond rearrangement causes a bond cleavage between the substrate and GSH.

Products.

Contributors

Marko Babić, Antonio Ribeiro