Hydroxyacylglutathione hydrolase

 

Glyoxalase II participates in the cellular detoxification of cytotoxic and mutagenic 2-oxoaldehydes. Glyoxalase I converts methylglyoxal (a cytotoxic byproduct of many cellular reactions) and glutathione into S-D-lactoylglutathione (SLG) which is also cytotoxic; the role of glyoxalase II is to hydrolyse SLG into D-lactic acid and glutathione.

The substrate of this reaction is the product of Lactoylglutathione lyase, an enzyme which catalyses the initial steps in detoxifying methyl-glyoxyl. This enzyme regnerates the glutathione cofactor utilised by lactoylglutathione lyase.

 

Reference Protein and Structure

Sequence
Q16775 UniProt (3.1.2.6) IPR017782 (Sequence Homologues) (PDB Homologues)
Biological species
Homo sapiens (Human) Uniprot
PDB
1qh5 - HUMAN GLYOXALASE II WITH S-(N-HYDROXY-N-BROMOPHENYLCARBAMOYL)GLUTATHIONE (1.45 Å) PDBe PDBsum 1qh5
Catalytic CATH Domains
3.60.15.10 CATHdb (see all for 1qh5)
Cofactors
Zinc(2+) (2) Metal MACiE
Click To Show Structure

Enzyme Reaction (EC:3.1.2.6)

water
CHEBI:15377ChEBI
+
(R)-S-lactoylglutathionate(1-)
CHEBI:57474ChEBI
→
(R)-lactate
CHEBI:16004ChEBI
+
glutathionate(1-)
CHEBI:57925ChEBI
+
hydron
CHEBI:15378ChEBI
Alternative enzyme names: S-2-hydroxylacylglutathione hydrolase, Acetoacetylglutathione hydrolase, Glyoxalase II,

Enzyme Mechanism

Introduction

Glyoxalase II has a binuclear centre that has affinity for both Zn(II) and Fe(II) and is functional with either metal in either of its metal-binding sites. Both metals coordinate a bridging hydroxide ion. This hydroxide nucleophile attacks the carbonyl carbon of the thioester group of SLG. A tetrahedral intermediate is formed. Metal 1 stabilises the negative charge forming on the thioester oxygen, and metal 2 stabilises the negative charge forming on the thioester sulphur (thus making the thiolate anion a better leaving group). The tetrahedral intermediate collapses. Lactic acid and glutathione (still bound to metal 2 as a thiolate) are formed. Asp58 accepts a proton from the bridging oxygen to generate the lactate, this proton is then transferred to the thiolate to form glutathione. The products are released and new water bridges the zinc ions, the water is deprotonated by Asp58 regenerating the active site.

Catalytic Residues Roles

UniProt PDB* (1qh5)
Asp106 Asp58A Deprotonates lactic acid and transfers the proton to reform glutathione. Also deprotonates a water molecule to regenerate the active site. hydrogen bond acceptor, metal ligand, proton acceptor, proton donor, activator, electrostatic stabiliser
Asp182 Asp134A Acts as a bridging ligand between the two zinc sites. metal ligand
His102, His104, His158 His54A, His56A, His110A Forms part of the zinc 1 binding site. metal ligand
His107, His221 His59A, His173A Forms part of the zinc 2 binding site. metal ligand
*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

overall reactant used, bimolecular nucleophilic addition, unimolecular elimination by the conjugate base, overall product formed, proton transfer, native state of enzyme regenerated, native state of cofactor regenerated

References

  1. Zang TM et al. (2001), J Biol Chem, 276, 4788-4795. Arabidopsis Glyoxalase II Contains a Zinc/Iron Binuclear Metal Center That Is Essential for Substrate Binding and Catalysis. DOI:10.1074/jbc.m005090200. PMID:11085979.
  2. Chen SL et al. (2009), J Inorg Biochem, 103, 274-281. Reaction mechanism of the binuclear zinc enzyme glyoxalase II - A theoretical study. DOI:10.1016/j.jinorgbio.2008.10.016. PMID:19062100.
  3. Cameron AD et al. (1999), Structure, 7, 1067-1078. Crystal structure of human glyoxalase II and its complex with a glutathione thiolester substrate analogue. DOI:10.1016/s0969-2126(99)80174-9. PMID:10508780.

Step 1. Zinc activate water attacked the carbonyl carbon of the (R)-S-lactoylglutathione substrate in a nucleophilic addition, forming a tetrahedral intermediate.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His54A metal ligand
His56A metal ligand
His110A metal ligand
Asp58A hydrogen bond acceptor, metal ligand, electrostatic stabiliser
Asp134A metal ligand
His59A metal ligand
His173A metal ligand

Chemical Components

overall reactant used, ingold: bimolecular nucleophilic addition

Step 2. The tetrahedral intermediate collapses releasing the (R)-lactate product and the thiolate form of glutathione. Asp58 accepts a proton from the bridging oxygen.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His54A metal ligand
His56A metal ligand
Asp58A metal ligand
His59A metal ligand
His110A metal ligand
Asp134A metal ligand
His173A metal ligand
Asp58A hydrogen bond acceptor, electrostatic stabiliser
Asp58A proton acceptor

Chemical Components

ingold: unimolecular elimination by the conjugate base, overall product formed, proton transfer

Step 3. The thiolate of glutathione deprotonates Asp58, regenerating the glutathione cofactor used in the initial formation of hydroacetylglutathione.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp58A hydrogen bond acceptor, activator
His54A metal ligand
His56A metal ligand
His110A metal ligand
Asp134A metal ligand
Asp58A metal ligand
His59A metal ligand
His173A metal ligand
Asp58A proton donor

Chemical Components

proton transfer, native state of enzyme regenerated

Step 4. Asp58 deprotonates a new bridging water molecule regenerating the active site. This proton can then be transferred from Asp58 to solution.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His54A metal ligand
His56A metal ligand
Asp58A metal ligand
His59A metal ligand
His110A metal ligand
Asp134A metal ligand
His173A metal ligand
Asp58A proton acceptor

Chemical Components

proton transfer, native state of cofactor regenerated
Download: Image, Marvin File

Step 1. Zinc activate water attacked the carbonyl carbon of the (R)-S-lactoylglutathione substrate in a nucleophilic addition, forming a tetrahedral intermediate.

Step 2. The tetrahedral intermediate collapses releasing the (R)-lactate product and the thiolate form of glutathione. Asp58 accepts a proton from the bridging oxygen.

Step 3. The thiolate of glutathione deprotonates Asp58, regenerating the glutathione cofactor used in the initial formation of hydroacetylglutathione.

Step 4. Asp58 deprotonates a new bridging water molecule regenerating the active site. This proton can then be transferred from Asp58 to solution.

Products.

Contributors

Gemma L. Holliday, Daniel E. Almonacid, Steven Smith, James Torrance, James Willey