Sulfite:Cytochrome c oxidoreductase from Thiobacillus novellus. Purification, characterization, and molecular biology of a heterodimeric member of the sulfite oxidase family.
Direct oxidation of sulfite to sulfate occurs in various photo- and chemotrophic sulfur oxidizing microorganisms as the final step in the oxidation of reduced sulfur compounds and is catalyzed by sulfite:cytochrome c oxidoreductase (EC ). Here we show that the enzyme from Thiobacillus novellus is a periplasmically located alphabeta heterodimer, consisting of a 40.6-kDa subunit containing a molybdenum cofactor and an 8.8-kDa mono-heme cytochrome c(552) subunit (midpoint redox potential, E(m8.0) = +280 mV). The organic component of the molybdenum cofactor was identified as molybdopterin contained in a 1:1 ratio to the Mo content of the enzyme. Electron paramagnetic resonance spectroscopy revealed the presence of a sulfite-inducible Mo(V) signal characteristic of sulfite:acceptor oxidoreductases. However, pH-dependent changes in the electron paramagnetic resonance signal were not detected. Kinetic studies showed that the enzyme exhibits a ping-pong mechanism involving two reactive sites. K(m) values for sulfite and cytochrome c(550) were determined to be 27 and 4 micrometer, respectively; the enzyme was found to be reversibly inhibited by sulfate and various buffer ions. The sorAB genes, which encode the enzyme, appear to form an operon, which is preceded by a putative extracytoplasmic function-type promoter and contains a hairpin loop termination structure downstream of sorB. While SorA exhibits significant similarities to known sequences of eukaryotic and bacterial sulfite:acceptor oxidoreductases, SorB does not appear to be closely related to any known c-type cytochromes.
Kappler
U
U
Institut für Mikrobiologie und Biotechnologie, Meckenheimer Allee 168, D-53115 Bonn, Germany.
Bennett
B
B
Rethmeier
J
J
Schwarz
G
G
Deutzmann
R
R
McEwan
A G
AG
Dahl
C
C
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D001483:Base Sequence; D003579:Cytochrome Reductases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D013379:Substrate Specificity; D050902:Sulfite Dehydrogenase; D013855:Thiobacillus / Q000201:enzymology*
D001426:Bacterial Proteins; D003579:Cytochrome Reductases; D050902:Sulfite Dehydrogenase