ABSTRACT Photosynthetic organisms have the unique ability to transform light energy into reducing power. We study the requirements for photosynthesis in the α-proteobacterium Rhodobacter sphaeroides . Global gene expression analysis found that ∼50 uncharacterized genes were regulated by changes in light intensity and O ₂ tension, similar to the expression of genes known to be required for photosynthetic growth of this bacterium. These uncharacterized genes included RSP4157 to -4159, which appeared to be cotranscribed and map to plasmid P004. A mutant containing a polar insertion in RSP4157, CT01, was able to grow via photosynthesis under autotrophic conditions using H ₂ as an electron donor and CO ₂ as a carbon source. However, CT01 was unable to grow photoheterotrophically in a succinate-based medium unless compounds that could be used to recycle reducing power (the external electron acceptor dimethyl sulfoxide (DMSO) or CO ₂ ) were provided. This suggests that the insertion in RSP4157 caused a defect in recycling reducing power during photosynthetic growth when a fixed carbon source was present. CT01 had decreased levels of RNA for genes encoding putative glycolate degradation functions. We found that exogenous glycolate also rescued photoheterotrophic growth of CT01, leading us to propose that CO ₂ produced from glycolate metabolism can be used by the Calvin cycle to recycle reducing power generated in the photosynthetic apparatus. The ability of glycolate, CO ₂ , or DMSO to support photoheterotrophic growth of CT01 suggests that one or more products of RSP4157 to -4159 serve a previously unknown role in recycling reducing power under photosynthetic conditions.