Biosynthesis of the respiratory formate dehydrogenases from Escherichia coli: characterization of the FdhE protein.
Escherichia coli can perform two modes of formate metabolism. Under respiratory conditions, two periplasmically-located formate dehydrogenase isoenzymes couple formate oxidation to the generation of a transmembrane electrochemical gradient; and under fermentative conditions a third cytoplasmic isoenzyme is involved in the disproportionation of formate to CO(2) and H(2). The respiratory formate dehydrogenases are redox enzymes that comprise three subunits: a molybdenum cofactor- and FeS cluster-containing catalytic subunit; an electron-transferring ferredoxin; and a membrane-integral cytochrome b. The catalytic subunit and its ferredoxin partner are targeted to the periplasm as a complex by the twin-arginine transport (Tat) pathway. Biosynthesis of these enzymes is under control of an accessory protein termed FdhE. In this study, it is shown that E. coli FdhE interacts with the catalytic subunits of the respiratory formate dehydrogenases. Purification of recombinant FdhE demonstrates the protein is an iron-binding rubredoxin that can adopt monomeric and homodimeric forms. Bacterial two-hybrid analysis suggests the homodimer form of FdhE is stabilized by anaerobiosis. Site-directed mutagenesis shows that conserved cysteine motifs are essential for the physiological activity of the FdhE protein and are also involved in iron ligation.
Lüke
Iris
I
Division of Molecular and Environmental Microbiology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, UK.
Butland
Gareth
G
Moore
Kevin
K
Buchanan
Grant
G
Lyall
Verity
V
Fairhurst
Shirley A
SA
Greenblatt
Jack F
JF
Emili
Andrew
A
Palmer
Tracy
T
Sargent
Frank
F
D020134:Catalytic Domain; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005560:Formate Dehydrogenases / Q000096:biosynthesis* / Q000737:chemistry; D033862:Iron-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry
D029968:Escherichia coli Proteins; C534169:FdhE protein, E coli; D033862:Iron-Binding Proteins; D026901:Membrane Transport Proteins; D011994:Recombinant Proteins; C408585:twin-arginine translocase complex, E coli; D005560:Formate Dehydrogenases; D003545:Cysteine