Abstract
Sulfate-reducing bacteria play major roles in the global carbon and sulfur cycles, but it remains unclear how reducing sulfate yields energy. To determine the genetic basis of energy conservation, we measured the fitness of thousands of pooled mutants of Desulfovibrio alaskensis G20 during growth in 12 different combinations of electron donors and acceptors. We show that ion pumping by the ferredoxin:NADH oxidoreductase Rnf is required whenever substrate-level phosphorylation is not possible. The uncharacterized complex Hdr/flox-1 (Dde_1207:13) is sometimes important alongside Rnf and may perform an electron bifurcation to generate more reduced ferredoxin from NADH to allow further ion pumping. Similarly, during the oxidation of malate or fumarate, the electron-bifurcating transhydrogenase NfnAB-2 (Dde_1250:1) is important and may generate reduced ferredoxin to allow additional ion pumping by Rnf. During formate oxidation, the periplasmic [NiFeSe] hydrogenase HysAB is required, which suggests that hydrogen forms in the periplasm, diffuses to the cytoplasm, and is used to reduce ferredoxin, thus providing a substrate for Rnf. During hydrogen utilization, the transmembrane electron transport complex Tmc is important and may move electrons from the periplasm into the cytoplasmic sulfite reduction pathway. Finally, mutants of many other putative electron carriers have no clear phenotype, which suggests that they are not important under our growth conditions, although we cannot rule out genetic redundancy.
Highlights
Sulfate-reducing bacteria are major players in the remineralization of fixed carbon and in the global sulfur cycle, but their energy metabolism remains poorly understood
Based on our genetic data, we propose an overview of electron flow and energy conservation in D. alaskensis G20 (Figure 1C) and specific scenarios of energy conservation with different electron donors (Figure 2; Presentation 1 in Supplementary Material)
GENOME-WIDE FITNESS DATA We used a collection of transposon mutants of D. alaskensis G20 that have been mapped and tagged with DNA barcodes (Kuehl et al, 2014)
Summary
Sulfate-reducing bacteria are major players in the remineralization of fixed carbon and in the global sulfur cycle, but their energy metabolism remains poorly understood. Research on the mechanism of sulfate reduction has focused on members of the genus Desulfovibrio, which are relatively easy to culture in the laboratory. Sulfate reduction is best studied in the strain Desulfovibrio vulgaris Hildenborough (Keller and Wall, 2011), but the Desulfovibrio genus is quite diverse. We are studying the energy metabolism of Desulfovibrio alaskensis G20 (formerly D. desulfuricans G20), for which a large collection of mutants is available (Kuehl et al, 2014). G20 is a derivative of the G100A strain that was isolated from an oil well in Ventura County, California (Wall et al, 1993). 1871 of 3258 proteins in the genome of D. alaskensis G20 (Hauser et al, 2011) have orthologs in D. vulgaris Hildenborough
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