The quinone-binding site of Acidithiobacillus ferrooxidans sulfide: quinone oxidoreductase controls both sulfide oxidation and quinone reduction.

Abstract

Sulfide:quinone oxidoreductase (SQR) is a peripheral membrane enzyme that catalyzes the oxidation of sulfide and the reduction of ubiquinone. Ubiquinone binds to a conserved hydrophobic domain and shuttles electrons from a noncovalent flavin adenine dinucleotide cofactor to the membrane-bound quinone pool. Utilizing the structure of decylubiquinone bound to Acidithiobacillus ferrooxidans SQR, we combined site-directed mutagenesis and kinetic approaches to analyze quinone binding. SQR can reduce both benzoquinones and naphthoquinones. The alkyl side-chain of ubiquinone derivatives enhances binding to SQR but limits the enzyme turnover. Pentachlorophenol and 2-n-heptyl-4-hydroxyquinoline-N-oxide are potent inhibitors of SQR with apparent inhibition constants (Ki) of 0.46 μmol·L(-1) and 0.58 μmol·L(-1), respectively. The highly conserved amino acids surrounding the quinone binding site play an important role in quinone reduction. The phenyl side-chains of Phe357 and Phe391 sandwich the benzoquinone head group and are critical for quinone binding. Importantly, conserved amino acids that define the ubiquinone-binding site also play an important role in sulfide oxidation/flavin reduction.