Methylglyoxal synthase

 

Methylglyoxyl synthase catalyses the first reaction in the methylglyoxyl bypass of the Embden-Myerhoff pathway (glycolysis), the conversion of dihydroxyacetone phosphate into methylglyoxal and orthophosphate. The physiological benefits of this are yet to be understood as the final product of methylglyoxal synthase is cytotoxic in small quantities and has been shown to be mutagenic and to interfere with de novo protein and nucleic acid synthesis. The intermediate, D-lactoylglutathione is also toxic in millimolar quantities, interfering with intermediate filament synthesis. The methylglyoxyl bypass system is inhibited allosterically by phosphate suggesting that is only functions in times of phosphate deprivation although more recent data imply that it may facilitate the transition between conditions of high and low phosphate. There is much interest in this enzyme for two reasons: firstly it has been implicated in diabetic complications and secondly due to its ability to metabolise anti-cancer drugs rendering treatments ineffective.

 

Reference Protein and Structure

Sequence
P0A731 UniProt (4.2.3.3) IPR004363 (Sequence Homologues) (PDB Homologues)
Biological species
Escherichia coli K-12 (Bacteria) Uniprot
PDB
1b93 - METHYLGLYOXAL SYNTHASE FROM ESCHERICHIA COLI (1.9 Å) PDBe PDBsum 1b93
Catalytic CATH Domains
3.40.50.1380 CATHdb (see all for 1b93)
Click To Show Structure

Enzyme Reaction (EC:4.2.3.3)

glycerone phosphate(2-)
CHEBI:57642ChEBI
→
hydrogenphosphate
CHEBI:43474ChEBI
+
methylglyoxal
CHEBI:17158ChEBI
Alternative enzyme names: Methylglyoxal synthetase, Glycerone-phosphate phospho-lyase,

Enzyme Mechanism

Introduction

The first step of the reaction mechanism involves the stereospecific abstraction of the C3 hydrogen of the substrate to form a common enediol enzyme intermediate. Asp71 abstracts a proton from the substrate whilst His98 protonates the ene-diol. His19 then abstracts the hydroxyl proton to initiate the phosphate elimination. Numerous other ligands stabilise intermediates formed in the active site.

Catalytic Residues Roles

UniProt PDB* (1b93)
Arg107 Arg107C Stabilises the negative charge on Asp101. hydrogen bond donor, electrostatic stabiliser
His19 His19A Acts as a general acid/base, activated by Asp91. hydrogen bond acceptor, hydrogen bond donor, proton acceptor, proton donor, electrostatic stabiliser
Asp91 Asp91A Activates the general acid/base His19. activator, hydrogen bond acceptor, electrostatic stabiliser
Asp71, His98 Asp71A, His98A Acts as a general acid/base. hydrogen bond acceptor, hydrogen bond donor, proton acceptor, proton donor
Asp101 Asp101A Activates Asp71 to act as the general acid/base through a repulsive charge-charge interaction. repulsive charge-charge interaction, activator, hydrogen bond acceptor, electrostatic stabiliser
Gly66 (main-N) Gly66A (main-N) The amide nitrogen of Gly66 positions and stabilises the bridging phosphoryl oxygen for bond cleavage. hydrogen bond donor, steric role, 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, assisted keto-enol tautomerisation, overall reactant used, intermediate formation, bimolecular elimination, dephosphorylation, intermediate collapse, overall product formed, intermediate terminated, native state of enzyme regenerated, inferred reaction step, reaction occurs outside the enzyme

References

  1. Saadat D et al. (2000), Biochemistry, 39, 2950-2960. Mirroring Perfection:  The Structure of Methylglyoxal Synthase Complexed with the Competitive Inhibitor 2-Phosphoglycolate†. DOI:10.1021/bi992666f. PMID:10715115.
  2. Marks GT et al. (2004), Biochemistry, 43, 3802-3813. Mutagenic Studies on Histidine 98 of Methylglyoxal Synthase:  Effects on Mechanism and Conformational Change†,‡. DOI:10.1021/bi035838o. PMID:15049687.
  3. Zhang X et al. (2002), J Am Chem Soc, 124, 14871-14878. Functional Specificities of Methylglyoxal Synthase and Triosephosphate Isomerase:  A Combined QM/MM Analysis. DOI:10.1021/ja027063x.
  4. Marks GT et al. (2001), Biochemistry, 40, 6805-6818. Mechanistic implications of methylglyoxal synthase complexed with phosphoglycolohydroxamic acid as observed by X-ray crystallography and NMR spectroscopy. PMID:11389594.
  5. Saadat D et al. (1999), Structure, 7, 309-317. The crystal structure of methylglyoxal synthase from Escherichia coli. DOI:10.1016/s0969-2126(99)80041-0. PMID:10368300.
  6. Saadat D et al. (1998), Biochemistry, 37, 10074-10086. Identification of Catalytic Bases in the Active Site ofEscherichia coliMethylglyoxal Synthase:  Cloning, Expression, and Functional Characterization of Conserved Aspartic Acid Residues†. DOI:10.1021/bi980409p. PMID:9665712.

Step 1. Asp71 deprotonates the terminal CH2, initiating a double bond rearrangement to form the enol-form of the substrate, with concomitant deprotonation of His98.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His19A hydrogen bond acceptor, hydrogen bond donor, electrostatic stabiliser
Gly66A (main-N) hydrogen bond donor
Asp71A hydrogen bond acceptor
Asp91A hydrogen bond acceptor
His98A hydrogen bond donor
Asp101A hydrogen bond acceptor, repulsive charge-charge interaction, activator
Arg107C hydrogen bond donor, electrostatic stabiliser
Asp71A proton acceptor
His98A proton donor

Chemical Components

proton transfer, assisted keto-enol tautomerisation, overall reactant used, intermediate formation

Step 2. His19 deprotonates the terminal alcohol group, initiating double bond rearrangement and the elimination of the phosphate group.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His19A hydrogen bond acceptor, hydrogen bond donor, electrostatic stabiliser
Gly66A (main-N) hydrogen bond donor, steric role, electrostatic stabiliser
Asp71A hydrogen bond donor
Asp91A hydrogen bond acceptor, activator
His98A hydrogen bond acceptor
Asp101A hydrogen bond acceptor, electrostatic stabiliser
Arg107C hydrogen bond donor, electrostatic stabiliser
His19A proton acceptor

Chemical Components

ingold: bimolecular elimination, dephosphorylation, intermediate formation, intermediate collapse, overall product formed

Step 3. His98 deprotonates His19 and the phosphate group is assumed to deprotonate Asp71 in an inferred step that returns the enzyme to its active state.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His19A hydrogen bond donor, electrostatic stabiliser
Gly66A (main-N) hydrogen bond donor, electrostatic stabiliser
Asp71A hydrogen bond donor
Asp91A hydrogen bond acceptor, electrostatic stabiliser
His98A hydrogen bond acceptor
Asp101A hydrogen bond acceptor, electrostatic stabiliser
Arg107C hydrogen bond donor, electrostatic stabiliser
His98A proton acceptor
Asp71A proton donor
His19A proton donor

Chemical Components

proton transfer, overall product formed, intermediate terminated, native state of enzyme regenerated, inferred reaction step

Step 4. The final tautomerisation occurs outside of the enzyme active site.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles

Chemical Components

reaction occurs outside the enzyme
Download: Image, Marvin File

Step 1. Asp71 deprotonates the terminal CH2, initiating a double bond rearrangement to form the enol-form of the substrate, with concomitant deprotonation of His98.

Step 2. His19 deprotonates the terminal alcohol group, initiating double bond rearrangement and the elimination of the phosphate group.

Step 3. His98 deprotonates His19 and the phosphate group is assumed to deprotonate Asp71 in an inferred step that returns the enzyme to its active state.

Step 4. The final tautomerisation occurs outside of the enzyme active site.

Products.

Contributors

Gemma L. Holliday, Gail J. Bartlett, Daniel E. Almonacid, Alex Gutteridge, Craig Porter