Formate dehydrogenase (nitrate-inducible)

 

Formate dehydrogenase-N (Fdn-N) isolated from Escherichia coli catalyses the oxidation of formate to carbon dioxide and a proton, coupled to the reduction of menaquinone to menaquinol. Fdn-N is involved in nitrate respiration and forms a redox loop with dissimilatory nitrate reductase (Nar). Together they pump protons across the membrane by reducing menaquinone at Fdn-N (taking up protons from the cytoplasm) and oxidising menaquinol at Nar, (releasing protons into the periplasm). They also generate a membrane potential by moving electrons from Fdn-N to Nar.

Fdn-N contains an active site for formate oxidation (alpha subunit) and an active site for menaquinone reduction (gamma subunit). They are linked by a redox chain containing two haem groups (gamma), five [4Fe-4S] clusters (FeS-0 in alpha and FeS-1 to FeS-4 in beta) and a molybdenum(VI) ion coordinated to two molybdopterin-guanine dinucleotides, the selenate group from Se-Cys196(alpha) and a hydroxide ion (alpha).

 

Reference Protein and Structure

Sequences
P24183 UniProt (1.17.5.3)
P0AAJ3 UniProt
P0AEK7 UniProt IPR006443 (Sequence Homologues) (PDB Homologues)
Biological species
Escherichia coli K-12 (Bacteria) Uniprot
PDB
1kqf - FORMATE DEHYDROGENASE N FROM E. COLI (1.6 Å) PDBe PDBsum 1kqf
Catalytic CATH Domains
3.40.228.10 CATHdb 1.20.950.20 CATHdb (see all for 1kqf)
Cofactors
Molybdopterin guanine dinucleotide (2), Molybdenum(6+) (1), Tetra-mu3-sulfido-tetrairon (5), Heme b (2)
Click To Show Structure

Enzyme Reaction (EC:1.17.5.3)

formate
CHEBI:15740ChEBI
+
menaquinone
CHEBI:16374ChEBI
+
hydron
CHEBI:15378ChEBI
→
carbon dioxide
CHEBI:16526ChEBI
+
menaquinol
CHEBI:18151ChEBI
Alternative enzyme names: Fdh-N, FdnGHI, Nitrate-inducible formate dehydrogenase, Formate dehydrogenase N, FDH-N, Nitrate inducible Fdn, Nitrate inducible formate dehydrogenase,

Enzyme Mechanism

Introduction

Formate displaces the selenate group of Se-Cys196(alpha) from the Mo(VI) centre. His197(alpha) accepts a proton from formate, which leads to the formation of carbon dioxide (possibly via a selenium-carboxylated intermediate) and the reduction of the molybdenum centre to Mo(IV). The selenate group then re-coordinates to the metal centre. An electron is passed down a chain of iron-sulfur centres in the order FeS-0, FeS-1, FeS-4, FeS-2 and then FeS-3. It is transferred to haem bp, haem bc and then His169(gamma). Menaquinone accepts the electron from His169(gamma) and a proton through a water chain to form menasemiquinone. Mo(V) donates another electron and menasemiquinone is reduced to menaquinol and accepts a proton from His169(gamma). Menaquinol is released and His169(gamma) is protonated through a water channel.

Catalytic Residues Roles

UniProt PDB* (1kqf)
Sec196 Sec196A The selenate group of Se-Cys196(alpha) is displaced from the molybdenum centre by formate. It is thought to be involved in a selenium-carboxylated intermediate before carbon dioxide is released. Once this has occurred the selenate can re-coordinate to the molybdenum centre. covalently attached, nucleofuge, metal ligand
His197 His197A His197(alpha) accepts a proton during the two-electron oxidation of formate.
His169 His169C(CA) His169(gamma) is a ligand for haem bc. It transfers electrons from the haem group to menaquinone/menasemiquinone. It also donates a proton to O-1 upon the reduction of menasemiquinone. It is reprotonated only after the menaquinol has been released and a water chain has formed. single electron relay, hydrogen bond donor, proton acceptor, proton donor, single electron acceptor, single electron donor
*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

coordination to a metal ion, decoordination from a metal ion, decarboxylation, proton transfer, electron transfer, electron relay, intermediate formation, overall product formed

References

  1. Jormakka M et al. (2002), Science, 295, 1863-1868. Molecular Basis of Proton Motive Force Generation: Structure of Formate Dehydrogenase-N. DOI:10.1126/science.1068186. PMID:11884747.
  2. Raaijmakers HC et al. (2006), J Biol Inorg Chem, 11, 849-854. Formate-reduced E. coli formate dehydrogenase H: the reinterpretation of the crystal structure suggests a new reaction mechanism. DOI:10.1007/s00775-006-0129-2. PMID:16830149.
  3. Blasco F et al. (2001), Cell Mol Life Sci, 58, 179-193. The coordination and function of the redox centres of the membrane-bound nitrate reductases. DOI:10.1007/pl00000846. PMID:11289300.

Step 1. Formate displaces Sec196 and coordinates to the Mo(VI) centre.

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Catalytic Residues Roles

Residue Roles
Sec196A covalently attached
Sec196A nucleofuge

Chemical Components

coordination to a metal ion, decoordination from a metal ion

Step 2. His197 accepts a proton from formate, forming carbon dioxide and reducing the Mo centre to Mo(IV). The proton abstraction may proceed via a selenium-carboxylated intermediate. Following this step Sec196 re-coordinates to the metal centre.

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Catalytic Residues Roles

Residue Roles
His169C(CA) proton acceptor

Chemical Components

decarboxylation, proton transfer, coordination to a metal ion, decoordination from a metal ion

Step 3. An electron is passed down a chain of iron-sulfur centres in the alpha then beta subunit of the protein.

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Catalytic Residues Roles

Residue Roles
Sec196A metal ligand

Chemical Components

electron transfer, electron relay

Step 4. The electron is then transferred to haem bp, haem bc, His169 then accepted by menaquinone in the gamma subunit of the protein. The oxygen accepts a proton through a water chain to form a radical menasemiquinone intermediate.The O1 atom of menaquinone accepts a hydrogen bond from His169.

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Catalytic Residues Roles

Residue Roles
His169C(CA) hydrogen bond donor, single electron relay, single electron donor, single electron acceptor

Chemical Components

electron relay, electron transfer, intermediate formation

Step 5. Mo(V) donates another electron which is transferred through the iron sulfur clusters and haem cofactors to menasemiquinone. Mo(V) is oxidised to regenerate Mo(VI) and menasemiquinone is reduced.

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Catalytic Residues Roles

Residue Roles
Sec196A metal ligand

Chemical Components

electron transfer, electron relay, electron transfer

Step 6. The alkoxide accepts a proton from His169 and menaquinol is released. Deprotonated His169 is very unstable so it is immediately reprotonated via an extended water channel.

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Catalytic Residues Roles

Residue Roles
His169C(CA) proton donor

Chemical Components

overall product formed, proton transfer
Download: Image, Marvin File

Step 1. Formate displaces Sec196 and coordinates to the Mo(VI) centre.

Step 2. His197 accepts a proton from formate, forming carbon dioxide and reducing the Mo centre to Mo(IV). The proton abstraction may proceed via a selenium-carboxylated intermediate. Following this step Sec196 re-coordinates to the metal centre.

Step 3. An electron is passed down a chain of iron-sulfur centres in the alpha then beta subunit of the protein.

Step 4. The electron is then transferred to haem bp, haem bc, His169 then accepted by menaquinone in the gamma subunit of the protein. The oxygen accepts a proton through a water chain to form a radical menasemiquinone intermediate.The O1 atom of menaquinone accepts a hydrogen bond from His169.

Step 5. Mo(V) donates another electron which is transferred through the iron sulfur clusters and haem cofactors to menasemiquinone. Mo(V) is oxidised to regenerate Mo(VI) and menasemiquinone is reduced.

Step 6. The alkoxide accepts a proton from His169 and menaquinol is released. Deprotonated His169 is very unstable so it is immediately reprotonated via an extended water channel.

Products.

Contributors

Gemma L. Holliday, Amelia Brasnett, Trung Nguyen