PMC

Deletion of tol1⁺ gene leads to sulfite auxotrophy. (A) Δtol1 cells (Δtol1) and Δtol1 cells carrying pDB248[tol1⁺] (Δtol1+ptol1⁺) or pDB248[HAL2] (Δtol1+pHAL2) were streaked onto each plate containing YPAD plus 16 mM sodium sulfite (+Sulfite) or YPAD (−Sulfite) and incubated at 33°C for 4 days. (B) Δtol1 cells carrying pDB248[tol1⁺] (Δtol1+ptol1⁺) or pREP81[tol1⁺] (Δtol1+pREPtol1⁺) were streaked onto each plate containing EMM (EMM), EMM plus 4 μM thiamine (+Thiamine), EMM plus 16 mM sodium sulfite (+Sulfite), EMM plus 4 μM thiamine and 16 mM sodium sulfite (+Thiamine +Sulfite), or EMM plus 4 μM thiamine and cysteine (30 μg/ml) (+Thiamine +Cysteine). The plates were incubated at 33°C for 4 days. Each EMM plate was supplemented with adenine sulfate (20 μg/ml).

Cytochrome maturation and heme lyase components are required for optimal sulfite reduction. Wildtype and mutant S. oneidensis were grown anaerobically with sulfite as the sole electron acceptor. Sulfite reduction was indicated by the production of hydrogen sulfide (H₂S). (a) Loss of either heme chaperone, SirG or CcmI, resulted in a lag in sulfite reduction, likely due to a reduction in functionally active reductase. Sulfite activity was abolished when both sirG and ccmI were deleted. (b) Sulfite reductase activity of the ΔsirGΔccmI double mutant was partially restored, similar to that of the respected single mutant, when complemented with either sirG or ccmI. (c) Loss of the heme lyase components SirH or SirEF resulted in decreased sulfite reduction. The ΔsirH mutant exhibited a 48-hour lag in sulfite reduction compared to wildtype, whereas the ΔsirEF mutant lacked any sulfite reduction. The ΔsirA mutant served as a negative control for sulfite reduction in all assays.

The schematic representation of sulfite pathway in plants (modified from Brychkova et al. ). Sulfite is produced from sulfate by two consecutive enzymatic reactions of ATP sulfurylase and APS reductase. The produced sulfite is further reduced to cysteine by sulfite reductase and O-acetylserine (thiol) lyse respectively, to UDP-sulfoquinovose by UDP-sulfoquinovose synthase and to thiosulfate by sulfurtransferases. Sulfite is also oxidized to sulfate by a molybdenum cofactor-containing enzyme, sulfite oxidase

Schematic representation of sulfite network enzymes in plants. Sulfite is generated from sulfate in two consecutive steps. In the first step, ATP sufurylase catalyzes the adenylation of sulfate to , and then sulfite is produced in the chloroplast by the glutathione-dependent . In the chloroplast, the generated sulfite can be further reduced to sulfide by the ferredoxin-dependent . The sulfide together with O-acetyl-l-Ser are the substrates for Cys biosynthesis catalyzed by . Alternatively, the chloroplast-localized sulfite can enter the sulfolipid reductive pathway to generate in two consecutive steps. In the first step, UDP-sulfoquinovase is catalyzed by , employing sulfite and UDP-Glc as substrates, while in the second step, is catalyzed by synthase, employing UDP-sulfoquinovose and diacylglycerols as substrates. Sulfite localized to the cytosol and mitochondria may be detoxified to thiosulfate by the or be generated by these enzymes from thiosulfate and cyanide (; ; ). Sulfite can be oxidized to sulfate by the molybdenum cofactor-containing enzyme, peroxisomal . Fd^ox, Oxidized ferredoxin; Fd^red, reduced ferredoxin; GSSG, oxidized glutathione; H₂O₂, hydrogen peroxide; PPi, diphosphate.