Mechanism of Sulfide-Quinone Reductase Investigated Using Site-Directed Mutagenesis and Sulfur Analysis^,

Abstract

Biological sulfide oxidation is a reaction occurring in all three domains of life. One enzyme responsible for this reaction in many bacteria has been identified as sulfide:quinone oxidoreductase (SQR). The enzyme from Rhodobacter capsulatus is a peripherally membrane-bound flavoprotein with a molecular mass of approximately 48 kDa, presumably acting as a homodimer. In this work, SQR from Rb. capsulatus has been modified with an N-terminal His tag and heterologously expressed in and purified from Escherichia coli. Three cysteine residues have been shown to be essential for the reductive half-reaction by site-directed mutagenesis. The catalytic activity has been nearly completely abolished after mutation of each of the cysteines to serine. A decrease in fluorescence on reduction by sulfide as observed for the wild-type enzyme has not been observed for any of the mutated enzymes. Mutation of a conserved valine residue to aspartate within the third flavin-binding domain led to a drastically reduced substrate affinity, for both sulfide and quinone. Two conserved histidine residues have been mutated individually to alanine. Both of the resulting enzymes exhibited a shift in the pH dependence of the SQR reaction. Polysulfide has been identified as a primary reaction product using spectroscopic and chromatographic methods. On the basis of these data, reaction mechanisms for sulfide-dependent reduction and quinone-dependent oxidation of the enzyme and for the formation of polysulfide are proposed.

Hydrogen Sulfide Is an Endogenous Potentiator of T Cell Activation

“…5, A and B). Steady state levels of H 2 S also depend on its rate of degradation via SQR, an O 2 -dependent mitochondrial enzyme (53,54). SQR was slightly decreased in activated versus resting T cells (Fig.…”

Purification and Characterization of Sulfide:Quinone Oxidoreductase from an Acidophilic Iron-Oxidizing Bacterium,Acidithiobacillus ferrooxidans

“…Furthermore, three SQR fingerprints containing three cystein residues essential for the reductive reaction 33) were highly conserved.…”

Sulfide-responsive transcriptional repressor SqrR functions as a master regulator of sulfide-dependent photosynthesis

“…In the case of SQRs, S 0 is conjugated to a variety of nucleophilic acceptors, such as sulfite (SO 3 2-), CN -, S 2-, or a lowmolecular-weight thiol, for example glutathione (22)(23)(24), the latter of which gives rise to glutathione persulfide. In the purple photosynthetic bacterium Rhodobacter capsulatus, SQR is the major enzyme responsible for catalyzing this first step in sulfide oxidation, with the level of SQR expression increasing after sulfide treatment (25). R. capsulatus can also grow photosynthetically, using sulfide as well as several inorganic and organic compounds as electron donors.…”