2-amino-4-hydroxy-6-hydroxymethyldihydropteridine diphosphokinase

 

6-Hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) catalyses the transfer of pyrophosphate from ATP to 6-hydroxymethyl-7,8-dihydropterin(HP), the first reaction in the folate biosynthesis.

Folate cofactors are essential for life - they are required for the biosynthesis of purine and pyrimidine bases and certain amino acids. Mammals obtain folate from their diets by an active transport system. In contrast, many microorganisms lack the active transport system and must synthesise folate de novo. Therefore, enzymes in the folate biosynthesis pathway, including HPPK, are target for developing antimicrobial agents.

 

Reference Protein and Structure

Sequence
P26281 UniProt (2.7.6.3) IPR000550 (Sequence Homologues) (PDB Homologues)
Biological species
Escherichia coli K-12 (Bacteria) Uniprot
PDB
1q0n - CRYSTAL STRUCTURE OF A TERNARY COMPLEX OF 6-HYDROXYMETHYL-7,8-DIHYDROPTERIN PYROPHOSPHOKINASE FROM E. COLI WITH MGAMPCPP AND 6-HYDROXYMETHYL-7,8-DIHYDROPTERIN AT 1.25 ANGSTROM RESOLUTION (1.25 Å) PDBe PDBsum 1q0n
Catalytic CATH Domains
3.30.70.560 CATHdb (see all for 1q0n)
Cofactors
Magnesium(2+) (2) Metal MACiE
Click To Show Structure

Enzyme Reaction (EC:2.7.6.3)

2-amino-6-(hydroxymethyl)-7,8-dihydropteridin-4-one
CHEBI:44841ChEBI
+
ATP(4-)
CHEBI:30616ChEBI
→
adenosine 5'-monophosphate(2-)
CHEBI:456215ChEBI
+
(7,8-dihydropterin-6-yl)methyl diphosphate(3-)
CHEBI:72950ChEBI
+
hydron
CHEBI:15378ChEBI
Alternative enzyme names: 2-amino-4-hydroxy-6-hydroxymethyldihydropteridine pyrophosphokinase, 7,8-dihydro-6-hydroxymethylpterin pyrophosphokinase, 7,8-dihydroxymethylpterin-pyrophosphokinase, H(2)-Pteridine-CH(2)OH pyrophosphokinase, HPPK, Hydroxymethyldihydropteridine pyrophosphokinase, 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase, 6-hydroxymethyl-7,8-dihydropterin diphosphokinase, 7,8-dihydro-6-hydroxymethylpterin diphosphokinase, ATP:2-amino-4-hydroxy-6-hydroxymethyl-7,8-dihydropteridine 6'-diphosphotransferase,

Enzyme Mechanism

Introduction

The catalysis follows an inline displacement mechanism. The hydroxyl group of HP performs a nucleophilic attack to the beta-phosphate group of ATP to yield HP-pyrophosphate and AMP. Arg92, Arg82 and two Mg(II) ions stabilise the transition state. Both Mg(II) ions activate the beta-phosphorus for the nucleophilic attack and the Mg(II) ion coordinated to the hydroxyl group of HP reduces the hydroxyl group's pKa to facilitate the reaction.

Catalytic Residues Roles

UniProt PDB* (1q0n)
Arg83 Arg82A Interacts with the alpha-phosphate of ATP to stabilise the negative charge developed in the transition state. hydrogen bond donor, electrostatic stabiliser
Asp96, Asp98 Asp95A, Asp97A Forms part of Mg(II) binding site, both residues bind one Mg(II) ion via their OD2 atoms, and the other Mg(II) ion via their OD1 atoms. metal ligand
Arg93 Arg92A Interacts with the alpha-, beta- or both phosphates of ATP to stabilise the negative charge developed in the transition state. hydrogen bond donor, 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, bimolecular nucleophilic substitution, overall reactant used, overall product formed

References

  1. Blaszczyk J et al. (2000), Structure, 8, 1049-1058. Catalytic Center Assembly of HPPK as Revealed by the Crystal Structure of a Ternary Complex at 1.25 Å Resolution. DOI:10.1016/s0969-2126(00)00502-5. PMID:11080626.
  2. Derrick JP (2008), Vitam Horm, 411-433. Chapter 15 The Structure and Mechanism of 6‐Hydroxymethyl‐7,8‐Dihydropterin Pyrophosphokinase. DOI:10.1016/S0083-6729(08)00415-9.
  3. Li Y et al. (2005), Biochemistry, 44, 8590-8599. Is the Critical Role of Loop 3 ofEscherichia coli6-Hydroxymethyl-7,8-dihydropterin Pyrophosphokinase in Catalysis Due to Loop-3 Residues Arginine-84 and Tryptophan-89? Site-Directed Mutagenesis, Biochemical, and Crystallographic Studies†,‡. DOI:10.1021/bi0503495. PMID:15952765.
  4. Li Y et al. (2003), Biochemistry, 42, 1581-1588. Catalytic Roles of Arginine Residues 82 and 92 ofEscherichia coli6-Hydroxymethyl-7,8-dihydropterin Pyrophosphokinase:  Site-Directed Mutagenesis and Biochemical Studies†. DOI:10.1021/bi026800z. PMID:12578371.

Step 1. Water deprotonates the alcohol group of 6-hydroxymethyl-7,8-dihydropterin, which in turn attacks the beta phosphate of the ATP in a nucelophilic substitution reaction that generates the 7,8-dihydro-6-(diphosphooxymethyl)pterin and AMP products

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp95A metal ligand
Asp97A metal ligand
Arg82A hydrogen bond donor, electrostatic stabiliser
Arg92A hydrogen bond donor, electrostatic stabiliser

Chemical Components

proton transfer, ingold: bimolecular nucleophilic substitution, overall reactant used, overall product formed
Download: Image, Marvin File

Step 1. Water deprotonates the alcohol group of 6-hydroxymethyl-7,8-dihydropterin, which in turn attacks the beta phosphate of the ATP in a nucelophilic substitution reaction that generates the 7,8-dihydro-6-(diphosphooxymethyl)pterin and AMP products

Products.

Contributors

Gemma L. Holliday, Daniel E. Almonacid, Mei Leung, Charity Hornby