Proteomic analysis of Clostridium thermocellum ATCC 27405 reveals the upregulation of an alternative transhydrogenase-malate pathway and nitrogen assimilation in cells grown on cellulose.

Clostridium thermocellum is a Gram-positive thermophilic anaerobic bacterium with the ability to directly convert cellulosic biomass into useful products such as ethanol and hydrogen. In this study, a quantitative comparative proteomic analysis of the organism was performed to identify proteins and biochemical pathways that are differentially utilized by the organism after growth on cellobiose or cellulose. The cytoplasmic and membrane proteomes of C. thermocellum grown on cellulose or cellobiose were quantitatively compared using a metabolic (15)N isotope labelling method in conjunction with nanoLC-ESI-MS/MS (liquid chromatography - electrospray ionization - tandem mass spectrometry). In total, 1255 proteins were identified in the study, and 129 of those were able to have their relative abundance per cell compared in at least one cellular compartment in response to the substrate provided. This study reveals that cells grown on cellulose increase their abundance of phosphoenolpyruvate carboxykinase while decreasing the abundance of pyruvate dikinase and oxaloacetate decarboxylase, suggesting that the organism diverts carbon flow into a transhydrogenase-malate pathway that can increase the production of the biosynthetic intermediates NADPH and GTP. Glutamate dehydrogenase was also found to have increased abundance in cellulose-grown cells, suggesting that the assimilation of ammonia is upregulated in cells grown on the cellulosic substrates. The results illustrate a mechanism by which C. thermocellum can divert carbon into alternative pathways for the purpose of producing biosynthetic intermediates necessary to respond to growth on cellulose, including transhydrogenation of NADH to NADPH and increased nitrogen assimilation.