Clostridium thermocellum ATCC27405 transcriptomic, metabolomic and proteomic profiles after ethanol stress

Abstract

BackgroundClostridium thermocellum is a candidate consolidated bioprocessing biocatalyst, which is a microorganism that expresses enzymes for both cellulose hydrolysis and its fermentation to produce fuels such as lignocellulosic ethanol. However, C. thermocellum is relatively sensitive to ethanol compared to ethanologenic microorganisms such as yeast and Zymomonas mobilis that are used in industrial fermentations but do not possess native enzymes for industrial cellulose hydrolysis.ResultsIn this study, C. thermocellum was grown to mid-exponential phase and then treated with ethanol to a final concentration of 3.9 g/L to investigate its physiological and regulatory responses to ethanol stress. Samples were taken pre-shock and 2, 12, 30, 60, 120, and 240 min post-shock, and from untreated control fermentations for systems biology analyses. Cell growth was arrested by ethanol supplementation with intracellular accumulation of carbon sources such as cellobiose, and sugar phosphates, including fructose-6-phosphate and glucose-6-phosphate. The largest response of C. thermocellum to ethanol shock treatment was in genes and proteins related to nitrogen uptake and metabolism, which is likely important for redirecting the cells physiology to overcome inhibition and allow growth to resume.ConclusionThis study suggests possible avenues for metabolic engineering and provides comprehensive, integrated systems biology datasets that will be useful for future metabolic modeling and strain development endeavors.

Toxicological challenges to microbial bioethanol production and strategies for improved tolerance

“…Proteomic membrane profiling in an ethanol tolerant C. thermocellum determined that 73 % of all membrane proteins, including those involved in carbohydrate transport and energy metabolism, were detected at lower levels relative to wild type strains (Williams et al 2007). Additional work by Yang et al (2012) employed an integrated transcriptomic, metabolomic, and proteomic approach to assess the physiological and regulatory responses of wild type cells challenged with ethanol in continuous cultures. Their analysis revealed that observed reductions in growth following ethanol stress correlated with inhibition of glycolysis and pyruvate catabolism, and the buildup of cellobiose and glycolytic intermediates.…”

The DOE Bioenergy Research Centers: History, Operations, and Scientific Output

“…A mutation in the alcohol dehydrogenase gene, adhE, conferred a significant ethanol tolerance phenotype to C. thermocellum [65][66][67]. Further biochemistry showed that one of the effects of this mutation was a change of redox co-factor specificity from NADH to NADPH.…”

Thermophilic hydrogen-producing bacteria inhabiting deep-sea hydrothermal environments represented by Caloranaerobacter

“…BLAST searches of the retrieved 16S rRNA gene sequences in GenBank indicated that most of the sequences were closely affiliated (90e99% similarity) with Caloranaerobacter azorensis MV1087, which was a thermophilic chemoorganotrophic bacterium isolated from a deep-sea hydrothermal chimney sample in the Mid-Atlantic Ridge [21]. The sequences clustering within the genus Clostridium were most closely related (90e92% similarity) to Clostridium thermocellum ATCC27405, which is a thermophilic, cellulolytic and ethanogenic bacterium capable of directly converting cellulosic substrate into ethanol [33]. Members of the genus Caminicella shared 99% identity with Caminicella sporogenes, which is a thermophilic, strictly chemo-organoheterotrophic bacterium isolated from a deepsea hydrothermal vent sample from the East-Pacific Rise [34].…”