PDBsum entry 2b7s

Oxidoreductase

PDB id: 2b7s

Contents

Protein chain

568 a.a. *

Ligands

HEM ×4

FAD

FUM

Metals

_NA

Waters ×773

* Residue conservation analysis

PDB id:

2b7s

Name:

Oxidoreductase

Title:

R381k mutant of flavocytochrome c3

Structure:

Fumarate reductase flavoprotein subunit. Chain: a. Synonym: flavocytochrome c3, flavocytochromE C, fcc3. Engineered: yes. Mutation: yes

Source:

Shewanella frigidimarina. Organism_taxid: 56812. Gene: fcc. Expressed in: shewanella frigidimarina. Expression_system_taxid: 56812.

Resolution:

2.12Å R-factor: 0.147 R-free: 0.197

Authors:

K.L.Pankhurst,C.G.Mowat,E.L.Rothery,C.S.Miles,M.D.Walkinshaw, G.A.Reid,S.K.Chapman

Key ref:

K.L.Pankhurst et al. (2006). A proton delivery pathway in the soluble fumarate reductase from Shewanella frigidimarina. J Biol Chem, 281, 20589-20597. PubMed id: 16699170 DOI: 10.1074/jbc.M603077200

Date:

05-Oct-05 Release date: 23-May-06

Protein chain

P0C278  (FCCA_SHEFR) -  Fumarate reductase (cytochrome) from Shewanella frigidimarina

Seq: 571 a.a.

Struc: 568 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.1.3.2.4 - fumarate reductase (cytochrome).
• Reaction: 2 Fe(III)-[cytochrome c] + succinate = fumarate + 2 Fe(II)-[cytochrome c] + 2 H⁺

2 × Fe(III)-[cytochrome c] + succinate (Bound ligand (Het Group name = FUM) corresponds exactly) = fumarate + 2 × Fe(II)-[cytochrome c] + 2 × H(+)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

Added reference

DOI no: 10.1074/jbc.M603077200

J Biol Chem 281:20589-20597 (2006)

PubMed id: 16699170

A proton delivery pathway in the soluble fumarate reductase from Shewanella frigidimarina.

K.L.Pankhurst, C.G.Mowat, E.L.Rothery, J.M.Hudson, A.K.Jones, C.S.Miles, M.D.Walkinshaw, F.A.Armstrong, G.A.Reid, S.K.Chapman.

ABSTRACT

The mechanism for fumarate reduction by the soluble fumarate reductase from Shewanella frigidimarina involves hydride transfer from FAD and proton transfer from the active-site acid, Arg-402. It has been proposed that Arg-402 forms part of a proton transfer pathway that also involves Glu-378 and Arg-381 but, unusually, does not involve any bound water molecules. To gain further insight into the importance of this proton pathway we have perturbed it by substituting Arg-381 by lysine and methionine and Glu-378 by aspartate. Although all the mutant enzymes retain measurable activities, there are orders-of-magnitude decreases in their k(cat) values compared with the wild-type enzyme. Solvent kinetic isotope effects show that proton transfer is rate-limiting in the wild-type and mutant enzymes. Proton inventories indicate that the proton pathway involves multiple exchangeable groups. Fast scan protein-film voltammetric studies on wild-type and R381K enzymes show that the proton transfer pathway delivers one proton per catalytic cycle and is not required for transporting the other proton, which transfers as a hydride from the reduced, protonated FAD. The crystal structures of E378D and R381M mutant enzymes have been determined to 1.7 and 2.1 A resolution, respectively. They allow an examination of the structural changes that disturb proton transport. Taken together, the results indicate that Arg-381, Glu-378, and Arg-402 form a proton pathway that is completely conserved throughout the fumarate reductase/succinate dehydrogenase family of enzymes.

Selected figure(s)

Figure 6.
FIGURE 6. Stereoview overlay of the wild-type (gray/atom-type colors), E378D (green), and R381K (purple) flavocytochromes c[3]. This figure was generated using BOBSCRIPT (39) and RASTER 3D (40).

Figure 7.
FIGURE 7. The reaction mechanism for fumarate reduction by flavocytochrome c[3] showing the pathways taken by each of the two protons required to complete the catalytic cycle. This is an abbreviated version of the mechanism previously reported (12). One proton is transferred directly (from solvent or at least by a pathway not involving Arg-381, Glu-378, or Arg-402) to the FAD in a reaction coupled to its two-electron reduction. The substrate is twisted out of plane and polarized by interactions with positively charged residues. This facilitates hydride transfer from flavin N5 to the fumarate C2. Arg-402 acts as the active site acid donating a proton to fumarate C3. Arg-402 is reprotonated via the proton pathway involving Glu-378 and Arg-381.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2006, 281, 20589-20597) copyright 2006.

Figures were selected by an automated process.