Non-growing Rhodopseudomonas palustris Increases the Hydrogen Gas Yield from Acetate by Shifting from the Glyoxylate Shunt to the Tricarboxylic Acid Cycle

When starved for nitrogen, non-growing cells of the photosynthetic bacterium Rhodopseudomonas palustris continue to metabolize acetate and produce H-2, an important industrial chemical and potential biofuel. The enzyme nitrogenase catalyzes H-2 formation. The highest H-2 yields are obtained when cells are deprived of N-2 and thus use available electrons to synthesize H-2 as the exclusive product of nitrogenase. To understand how R. palustris responds metabolically to increase H-2 yields when it is starved for N-2, and thus not growing, we tracked changes in biomass composition and global transcript levels. In addition to a 3.5-fold higher H-2 yield by non-growing cells we also observed an accumulation of polyhydroxybutyrate to over 30% of the dry cell weight. The transcriptome of R. palustris showed down-regulation of biosynthetic processes and up-regulation of nitrogen scavenging mechanisms in response to N-2 starvation but gene expression changes did not point to metabolic activities that could generate the reductant necessary to explain the high H-2 yield. We therefore tracked C-13-labeled acetate through central metabolic pathways. We found that non-growing cells shifted their metabolism to use the tricarboxylic acid cycle to metabolize acetate in contrast to growing cells, which used the glyoxylate cycle exclusively. This shift enabled cells to more fully oxidize acetate, providing the necessary reducing power to explain the high H-2 yield.