2B7R

Structure of E378D mutant flavocytochrome c3

Summary for 2B7R

• Entry DOI: 10.2210/pdb2b7r/pdb
• Related: 1QJD 2B7S
• Descriptor: Fumarate reductase flavoprotein subunit, SODIUM ION, PROTOPORPHYRIN IX CONTAINING FE, ... (6 entities in total)
• Functional Keywords: flavocytochrome c3, fumarate reductase, proton delivery, oxidoreductase
• Biological source: Shewanella frigidimarina
• Cellular location: Periplasm: Q02469
• Total number of polymer chains: 1
• Total formula weight: 64003.78

Authors

Pankhurst, K.L.,Mowat, C.G.,Rothery, E.L.,Miles, C.S.,Walkinshaw, M.D.,Reid, G.A.,Chapman, S.K. (deposition date: 2005-10-05, release date: 2006-05-23, Last modification date: 2024-10-16)

Primary citation

Pankhurst, K.L.,Mowat, C.G.,Rothery, E.L.,Hudson, J.M.,Jones, A.K.,Miles, C.S.,Walkinshaw, M.D.,Armstrong, F.A.,Reid, G.A.,Chapman, S.K.
A Proton Delivery Pathway in the Soluble Fumarate Reductase from Shewanella frigidimarina.
J.Biol.Chem., 281:20589-20597, 2006

PubMed 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.

PubMed: 16699170
DOI: 10.1074/jbc.M603077200

Experimental method

X-RAY DIFFRACTION (1.7 Å)