Fumarate reductase (quinol)

 

Fumarate respiration is the most commonly occurring type of anaerobic respiration with fumarate acting as a terminal electron acceptor. The physiological reductant is unknown, but evidence indicates that flavocytochrome c participates in electron transfer from formate to fumarate and possibly also to trimethylamine oxide (TMAO).

In Shewanella species, fumarate reductase is a soluble periplasmic, tetaheme, FAD-containing enzyme called flavocytochrome c3 (Fcc3).

Fumarate reductases, which catalyse the interconversion of fumarate and succinate are known to be membrane bound in bacteria although soluble versions also exist in yeast, procyclic Trypanosoma brucei and several Shewanella species. The active site is located in the centre of the protein, at the interface between the three catalytic domains. Catalysis in the soluble fumarate reductase is essentially unidirectional (from fumarte to succinate).

 

Reference Protein and Structure

Sequence
Q07WU7 UniProt (1.3.5.4) IPR010960 (Sequence Homologues) (PDB Homologues)
Biological species
Shewanella frigidimarina NCIMB 400 (Bacteria) Uniprot
PDB
1qjd - Flavocytochrome C3 from Shewanella frigidimarina (1.8 Å) PDBe PDBsum 1qjd
Catalytic CATH Domains
3.50.50.60 CATHdb 3.90.700.10 CATHdb (see all for 1qjd)
Cofactors
Ferroheme c(2-) (4), Fadh2(2-) (1) Metal MACiE
Click To Show Structure

Enzyme Reaction (EC:1.3.5.4)

fumarate(2-)
CHEBI:29806ChEBI
+
menaquinol
CHEBI:18151ChEBI
→
succinate(2-)
CHEBI:30031ChEBI
+
menaquinone
CHEBI:16374ChEBI
Alternative enzyme names: FRD, Menaquinol-fumarate oxidoreductase, Succinate dehydrogenase (menaquinone), Succinate:menaquinone oxidoreductase, Fumarate reductase (menaquinone), Complex II,

Enzyme Mechanism

Introduction

On binding at the active site, the C1 carboxylate group of the fumarate substrate is polarised and twisted out of the planar configuration by closure of the clamp domain and the resulting steric constraints.

The substrate carbonyl groups are polarised through interactions with surrounding charged residues, facilitating hydride transfer from N5 of reduced FAD to the substrate C2.

The polar hydrogen bonding environment of the C4 carboxylate, as a result of interactions with His504, Arg544 and Arg402, polarises the C2-C3 bond of the substrate, causing C2 to become increasingly susceptible to nucleophilic attack and facilitating hydride transfer from N5 of the reduced flavin to the si face of the substrate. Concurrently, a proton is transferred to the substrate from His504.

Arg402 acts as an acid catalyst, transferring a proton to the substrate as part of a proton delivery pathway involving Arg381 and Glu378, resulting in the formation of the product, succinate.

Catalytic Residues Roles

UniProt PDB* (1qjd)
His529 His504A During the hydride transfer from the FAD cofactor to the fumarate C1, the residue stabilises the forming anionic charge on the oxygen through hydrogen bonding and subsequent proton donation. The residue is activated towards its catalytic role through interactions with His505. proton acceptor, electrostatic stabiliser, proton donor
His530 His505A His 505 modifies the pKa of the general acid His504 through hydrogen bond interactions, activating the residue towards its catalytic role. electrostatic stabiliser
Arg569 Arg544A Electrostatic interactions between the anionic carboxylate group and the residue's cationic side chain twist the substrate into a reactive conformation and polarise the carboxylate carbonyl which facilitates hydride transfer to the double bond C3. steric role
Arg406 Arg381A Part of the proton relay from bulk solvent to the general acid Arg402. proton relay, proton acceptor, proton donor
His390 His365A Activates water for the initial hydride transfer to occur. increase acidity
Glu403 Glu378A The residue relays a proton to the general acid Arg402 from Arg381. proton relay, proton acceptor, proton donor
Arg427 Arg402A Once hydride transfer has occurred, the residue is correctly positioned to donate a proton to the substrate C3 position, resulting in the formation of succinate. Arg402 transfers protons to the substrate as part of a proton delivery pathway involving Arg381 and Glu378. The residue is also acting as a Lewis acid, stabilising the build-up of negative charge in the transition state. proton acceptor, proton 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

electron relay, proton transfer, hydride transfer, aromatic unimolecular elimination by the conjugate base, bimolecular nucleophilic addition, assisted keto-enol tautomerisation

References

  1. Reid GA et al. (2000), Biochim Biophys Acta, 1459, 310-315. Catalysis in fumarate reductase. DOI:10.1016/s0005-2728(00)00166-3. PMID:11004445.
  2. Guillén Schlippe YV et al. (2005), Arch Biochem Biophys, 433, 266-278. A twisted base? The role of arginine in enzyme-catalyzed proton abstractions. DOI:10.1016/j.abb.2004.09.018. PMID:15581582.
  3. Schwalb C et al. (2003), Biochemistry, 42, 9491-9497. The Tetraheme Cytochrome CymA Is Required for Anaerobic Respiration with Dimethyl Sulfoxide and Nitrite inShewanella oneidensis†. DOI:10.1021/bi034456f. PMID:12899636.
  4. Mowat CG et al. (2002), Biochemistry, 41, 11990-11996. Engineering Water To Act as an Active Site Acid Catalyst in a Soluble Fumarate Reductase†. DOI:10.1021/bi0203177. PMID:12356299.
  5. Pankhurst KL et al. (2002), Biochemistry, 41, 8551-8556. Role of His505 in the Soluble Fumarate Reductase fromShewanella frigidimarina†. DOI:10.1021/bi020155e. PMID:12093271.
  6. Jeuken LJ et al. (2002), J Am Chem Soc, 124, 5702-5713. Electron-Transfer Mechanisms through Biological Redox Chains in Multicenter Enzymes. DOI:10.1021/ja012638w. PMID:12010043.
  7. Mowat CG et al. (2001), Biochemistry, 40, 12292-12298. Kinetic and Crystallographic Analysis of the Key Active Site Acid/Base Arginine in a Soluble Fumarate Reductase†. DOI:10.1021/bi011360h. PMID:11591148.
  8. Doherty MK et al. (2000), Biochemistry, 39, 10695-10701. Identification of the Active Site Acid/Base Catalyst in a Bacterial Fumarate Reductase:  A Kinetic and Crystallographic Study†. DOI:10.1021/bi000871l. PMID:10978153.
  9. Taylor P et al. (1999), Nat Struct Biol, 6, 1108-1112. Structural and mechanistic mapping of a unique fumarate reductase. DOI:10.1038/70045. PMID:10581550.
  10. Leys D et al. (1999), Nat Struct Biol, 6, 1113-1117. Structure and mechanism of the flavocytochrome c fumarate reductase of Shewanella putrefaciens MR-1. DOI:10.1038/70051. PMID:10581551.
  11. Bamford V et al. (1999), Nat Struct Biol, 6, 1104-1107. Open conformation of a flavocytochrome c3 fumarate reductase. DOI:10.1038/70039. PMID:10581549.

Step 1. The electron acceptors donates two electrons, through a chain of haem cofactors to the FAD cofactor, which deprotonates a base, probably water.

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Catalytic Residues Roles

Residue Roles
His365A increase acidity

Chemical Components

electron relay, proton transfer

Step 2. FAD eliminates a hydride ion, which adds to fumarate with concomitant double bond rearrangement, the terminal carboxylate deprotonates His504.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His504A electrostatic stabiliser
His505A electrostatic stabiliser
Arg544A steric role
His504A proton donor

Chemical Components

proton transfer, hydride transfer, ingold: aromatic unimolecular elimination by the conjugate base, ingold: bimolecular nucleophilic addition

Step 3. His504 deprotonates the newly formed hydroxyl group with concomitant deprotonation of Arg402, across the C=C.

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Catalytic Residues Roles

Residue Roles
Arg544A steric role
Arg402A proton donor
His504A proton acceptor

Chemical Components

proton transfer, assisted keto-enol tautomerisation

Step 4. A proton relay chain through Glu378 and Arg381 to bulk solvent reprotonates Arg402.

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Catalytic Residues Roles

Residue Roles
Arg381A proton donor
Arg402A proton acceptor
Arg381A proton acceptor
Glu378A proton donor, proton acceptor, proton relay
Arg381A proton relay

Chemical Components

proton transfer
Download: Image, Marvin File

Step 1. The electron acceptors donates two electrons, through a chain of haem cofactors to the FAD cofactor, which deprotonates a base, probably water.

Step 2. FAD eliminates a hydride ion, which adds to fumarate with concomitant double bond rearrangement, the terminal carboxylate deprotonates His504.

Step 3. His504 deprotonates the newly formed hydroxyl group with concomitant deprotonation of Arg402, across the C=C.

Step 4. A proton relay chain through Glu378 and Arg381 to bulk solvent reprotonates Arg402.

Products.

Contributors

Gemma L. Holliday, Daniel E. Almonacid, Gail J. Bartlett, Sophie T. Williams, Craig Porter, Katherine Ferris, Craig Porter