Phosphonate dehydrogenase

 

Phosphonate dehydrogenase, originally isolated from the Gram-negative bacterium Pseudomonas stutzeri (strain WM88), was the first protein known to catalyze redox chemistry on inorganic phosphorus compounds. Phosphonate dehydrogenase catalyzes the oxidation of phosphonate to phosphate with the concurrent reduction of NAD+ to NADH. Due to the strong exergonic nature of the reaction, phosphonate dehydrogenase has gained a particularly high interest for use as a cofactor regeneration enzyme.

 

Reference Protein and Structure

Sequence
O69054 UniProt (1.20.1.1) IPR006139 (Sequence Homologues) (PDB Homologues)
Biological species
Pseudomonas stutzeri (Bacteria) Uniprot
PDB
4e5k - Thermostable phosphite dehydrogenase in complex with NAD and sulfite (1.95 Å) PDBe PDBsum 4e5k
Catalytic CATH Domains
3.40.50.720 CATHdb (see all for 4e5k)
Click To Show Structure

Enzyme Reaction (EC:1.20.1.1)

water
CHEBI:15377ChEBI
+
NAD(1-)
CHEBI:57540ChEBI
+
phosphonate(2-)
CHEBI:16215ChEBI
→
hydron
CHEBI:15378ChEBI
+
NADH(2-)
CHEBI:57945ChEBI
+
hydrogenphosphate
CHEBI:43474ChEBI
Alternative enzyme names: NAD:phosphite oxidoreductase, Phosphite dehydrogenase, NAD-dependent phosphite dehydrogenas,

Enzyme Mechanism

Introduction

The catalytic mechanism of phosphonate dehydrogenase includes deprotonation of a water molecule by His292, followed by nucleophilic attack of resulting hydroxide on phosphonate together with the direct transfer of a hydride leaving group to NAD+, forming NADH and phosphate product. Met53 plays a catalytic role through transient stabilization of a positively charged His292 side chain.

Catalytic Residues Roles

UniProt PDB* (4e5k)
Met53 Met53B His53 plays a catalytic role through transient stabilization of a positively charged histidine side chain. electrostatic stabiliser
His292 His292B His292 acts as a general acid/base and abstracts proton from a water molecule for subsequent nucleophilic addition steps proton acceptor, proton donor
Arg237 Arg237B Arg237 is important for substrate binding and stabilization of the negative charges on the phosphonate and phosphate. 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 addition, aromatic bimolecular nucleophilic addition, overall product formed, overall reactant used

References

  1. Ranaghan KE et al. (2014), Chem Sci, 5, 2191-2199. A catalytic role for methionine revealed by a combination of computation and experiments on phosphite dehydrogenase. DOI:10.1039/c3sc53009d.
  2. Vrtis JM et al. (2001), J Am Chem Soc, 123, 2672-2673. Phosphite dehydrogenase: an unusual phosphoryl transfer reaction. DOI:10.1021/ja004301k. PMID:11456941.
  3. Costas AM et al. (2001), J Biol Chem, 276, 17429-17436. Purification and characterization of a novel phosphorus-oxidizing enzyme from Pseudomonas stutzeri WM88. DOI:10.1074/jbc.M011764200. PMID:11278981.

Step 1. Deprotonation of water molecule by His292, followed by nucleophilic attack of resulting hydroxide on phosphonate.

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Catalytic Residues Roles

Residue Roles
His292B proton acceptor
Arg237B electrostatic stabiliser

Chemical Components

proton transfer, ingold: bimolecular nucleophilic addition

Step 2. Nucleophilic addition of hydride leaving group to NAD+, forming NADH and phosphate product. Met53 plays a catalytic role through transient stabilization of a positively charged His292 side chain.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Met53B electrostatic stabiliser
Arg237B electrostatic stabiliser

Chemical Components

ingold: aromatic bimolecular nucleophilic addition, overall product formed, overall reactant used

Step 3. In an inferred reaction step His292 is deprotonated to regenerate the native state of the enzyme.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His292B proton donor
Arg237B electrostatic stabiliser

Chemical Components

proton transfer
Download: Image, Marvin File

Step 1. Deprotonation of water molecule by His292, followed by nucleophilic attack of resulting hydroxide on phosphonate.

Step 2. Nucleophilic addition of hydride leaving group to NAD+, forming NADH and phosphate product. Met53 plays a catalytic role through transient stabilization of a positively charged His292 side chain.

Step 3. In an inferred reaction step His292 is deprotonated to regenerate the native state of the enzyme.

Products.

Contributors

Trung Nguyen, Antonio Ribeiro