Rhizobia are nitrogen fixing bacteria that engage in symbiotic relationships with plant hosts but can also persist as free-living bacteria with the soil and rhizosphere. Here we show that free living Rhizobium leguminosarum SRDI565 can grow on the sulfosugar sulfoquinovose (SQ) using a sulfoglycolytic Entner-Doudoroff (sulfo-ED) pathway resulting in production of sulfolactate (SL) as the major metabolic end-product. Comparative proteomics supports the involvement of a sulfo-ED operon encoding

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... ABC transporter cassette, sulfo-ED enzymes and an SL exporter. Consistent with an oligotrophic lifestyle, proteomics data revealed little change in expression of the sulfo-ED proteins during growth on SQ versus mannitol, a result confirmed through biochemical assay of sulfoquinovosidase activity in cell lysates (data are available via ProteomeXchange with identifier PXD015822). Metabolomics analysis showed that growth on SQ involves gluconeogenesis to satisfy metabolic requirements for glucose-6-phosphate and fructose-6-phosphate. Metabolomics analysis also revealed the unexpected production of small amounts of sulfofructose and 2,3-dihydroxypropanesulfonate, which are proposed to arise from promiscuous activities of the glycolytic enzyme phosphoglucose isomerase and a promiscuous aldehyde reductase, respectively. This work shows that rhizobial metabolism of the abundant sulfosugar SQ may contribute to the mobilization of sulfur in soils.