title: Biochemical similarities and differences between the catalytic [4Fe-4S] cluster containing fumarases FumA and FumB from Escherichia coli.
abstract: BACKGROUND
  The highly homologous [4Fe-4S] containing fumarases FumA and FumB, sharing 90% amino acid sequence identity, from Escherichia coli are differentially regulated, which suggests a difference in their physiological function. The ratio of FumB over FumA expression levels increases by one to two orders of magnitude upon change from aerobic to anaerobic growth conditions.
  
  
  RESULTS
  To understand this difference in terms of structure-function relations, catalytic and thermodynamic properties were determined for the two enzymes obtained from homologous overexpression systems. FumA and FumB are essentially identical in their Michaelis-Menten kinetics of the reversible fumarate to L-malate conversion; however, FumB has a significantly greater catalytic efficiency for the conversion of D-tartrate to oxaloacetate consistent with the requirement of the fumB gene for growth on D-tartrate. Reduction potentials of the [4Fe-4S](2+) Lewis acid active centre were determined in mediated bulk titrations in the presence of added substrate and were found to be approximately -290 mV for both FumA and FumB.
  
  
  CONCLUSIONS
  This study contradicts previously published claims that FumA and FumB exhibit different catalytic preferences for the natural substrates L-malate and fumarate. FumA and FumB differ significantly only in the catalytic efficiency for the conversion of D-tartrate, a supposedly non-natural substrate. The reduction potential of the substrate-bound [4Fe-4S] active centre is, contrary to previously reported values, close to the cellular redox potential.
authors: Department of Biotechnology, Delft University of Technology, Delft, The Netherlands.
authors: Barbara M A
authors: BM
authors: van Vugt-Lussenburg
authors: Laura
authors: L
authors: van der Weel
authors: Wilfred R
authors: WR
authors: Hagen
authors: Peter-Leon
authors: PL
authors: Hagedoorn
mesh_terms: D000595:Amino Acid Sequence; D002384:Catalysis; D004578:Electron Spin Resonance Spectroscopy; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology*; D005649:Fumarate Hydratase / Q000737:chemistry / Q000378:metabolism*; D007501:Iron / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000737:chemistry / Q000378:metabolism*; D007527:Isoenzymes; D007700:Kinetics; D008969:Molecular Sequence Data; D062907:Oxaloacetic Acid / Q000378:metabolism; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013455:Sulfur / Q000378:metabolism*; D013644:Tartrates / Q000378:metabolism
chemical_list: D007506:Iron-Sulfur Proteins; D007527:Isoenzymes; D013644:Tartrates; D062907:Oxaloacetic Acid; D013455:Sulfur; D007501:Iron; D005649:Fumarate Hydratase; D010100:Oxygen; C029768:tartaric acid