Respiratory enzymes of Thiobacillus ferrooxidans. Kinetic properties of an acid-stable iron:rusticyanin oxidoreductase.
Rusticyanin is an acid-stable, soluble blue copper protein found in abundance in the periplasmic space of Thiobacillus ferrooxidans, an acidophilic bacterium capable of growing autotrophically on soluble ferrous sulfate. An acid-stable iron:rusticyanin oxidoreductase activity was partially purified from cell-free extracts of T. ferrooxidans. The enzyme-catalyzed, iron-dependent reduction of the rusticyanin exhibited three kinetic properties characteristic of aerobic iron oxidation by whole cells. (i) A survey of 14 different anions indicated that catalysis by the oxidoreductase occurred only in the presence of sulfate or selenate, an anion specificity identical to that of whole cells. (ii) Saturation with both sulfatoiron(II) and the catalyst produced a concentration-independent rate constant of 3 s-1 for the reduction of the rusticyanin, which is an electron transfer reaction sufficiently rapid to account for the flux of electrons through the iron respiratory chain. (iii) Values for the enzyme-catalyzed pseudo-first-order rate constants for the reduction of the rusticyanin showed a hyperbolic dependence on the concentration of sulfatoiron(II) with a half-maximal effect at 300 microM, a value similar to the apparent KM for iron shown by whole cells. On the basis of these favorable comparisons between the behavior patterns of isolated biomolecules and those of whole cells, this iron:rusticyanin oxidoreductase is postulated to be the primary cellular oxidant of ferrous ions in the iron respiratory electron transport chain of T. ferrooxidans.
Blake
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RC
Department of Biochemistry, Meharry Medical College, Nashville, Tennessee 37208.
Shute
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EA
D013856:Acidithiobacillus thiooxidans / Q000201:enzymology*; D000332:Aerobiosis; D001400:Azurin / Q000031:analogs & derivatives / Q000302:isolation & purification / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D003580:Cytochromes / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D007700:Kinetics; D008962:Models, Theoretical; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism*; D010101:Oxygen Consumption; D013053:Spectrophotometry
D001426:Bacterial Proteins; D003580:Cytochromes; C012543:rusticyanin; D001400:Azurin; D010088:Oxidoreductases; C088726:iron-rusticyanin oxidoreductase