Project description:Clostridium acetobutylicum is a typical bacterium of major importance to industrial butanol production. In order to dissect the regulatory network pertaining to the industrial application of this bacterium, catabolite control protein A (CcpA) was investigated for its global function by DNA microarray.It showed that CcpA of C. acetobutylicum controls hundreds of genes, not only carbon metabolism, but also solvent production and sporulation in the life cycle.The results here demonstrated that CcpA is an important pleiotropic regulator related to some specific physiological and biochemical process in butanol-producing C. acetobutylicum.
Project description:In this study the transcriptional behavior of the natural solvent producing bacterium Clostridium acetobutylicum was investigated following n-butanol stress using DNA microarray analysis. Therefore, a phosphate-limited chemostat culture was established and n-butanol stress (0.9%) was added to acidogenic cells at pH 5.7.
Project description:In this study the transcriptional behavior of the natural solvent producing bacterium Clostridium acetobutylicum was investigated following n butanol stress using DNA microarray analysis. Therefore, a phosphate-limited chemostat culture was established and n-butanol stress (0.9%) was added to acidogenic cells at pH 5.7.
Project description:Clostridium acetobutylicum is a typical bacterium of major importance to industrial butanol production. In order to dissect the regulatory network pertaining to the industrial application of this bacterium, catabolite control protein A (CcpA) was investigated for its global function by DNA microarray.It showed that CcpA of C. acetobutylicum controls hundreds of genes, not only carbon metabolism, but also solvent production and sporulation in the life cycle.The results here demonstrated that CcpA is an important pleiotropic regulator related to some specific physiological and biochemical process in butanol-producing C. acetobutylicum. In order to enable a global understanding of the regulatory roles of CcpA when fermenting mixed sugars, which is of great significance in utilization of lignocellulosic hydrolysates, D-glucose plus D-xylose were used as the carbon sources in fermentation for microarray analysis. Microarray analysis was performed at four time points:the time point M and L were chosen both in acidogenic phase, while the time point T and S were chosen in shift phase (from acidogenesis to solventogenesis) and solventogenic phase, respectively.One-color microarray assays were performed.Raw data were normalized by Quantile algorithm, Gene Spring Software 11.0 (Agilent technologies, Santa Clara, CA, US). The ratio of transcript level between wildtype and mutant can been achieved using the formula: 2^(value of wildtype)/2^(value of ccpA mutant).
Project description:Clostridium acetobutylicum is well-known for its butanol production. Butanol toxicity is a major drawback for the generation of high-butanol producing strains. Here, the transcriptional response a steady state, acidogenic (pH 6), phosphate-limited Clostridium acetobutylicum chemostat culture to different levels of n-butanol (0.25-1%) was investigated. For the butanol challenge experiments butanol (1-butanol) was added (a) to the supplying medium and (b) to the culture vessel to guarantee an immediate change in the butanol concentration. Addition of butanol to the culture was timed to match the supply of the new medium through the feedlines. The butanol concentration was increased stepwise in intervals of 66.6 h (5 volume changes) to moderate butanol concentrations of 0.25%, 0.5%, 0.75% and 1% (v/v).
Project description:Metabolite accumulation has pleiotropic, including toxic, effects on cellular physiology, with a variety of genetic resistance mechanisms previously identified. Using random genomic libraries and DNA microarrays, library inserts were preferentially enriched by culturing in media containing butanol, with successive transfers into fresh media containing incrementally increasing butanol concentrations. Keywords: genomic library preferential enrichment
Project description:Clostridium acetobutylicum is a Gram positive, endospore forming firmicute that has been known as the model organims for ABE (acetone-butanol-ethanol) fermentation. With its ability to consume a wide variety of substrates, C. acetobutylicum carries out a biphasic ABE fermentation, which consists of the acidogenic growth phase with the formation of butyric acid and acetic acid, followed by the solventogenic stationary phase with the formation of acetone, butanol and ethanol, characterised by the reassimilation of acids. The production butanol is of renewed ineterest both as a potential biofuel and bulk chemical production. Both butanol and butyrate posses toxic characteristic and here, we focus on understanding and modeling the stress response of C. acetobutylicum to one of the two important toxic metabolites: butanol.
Project description:Clostridium acetobutylicum is a Gram-positive, endospore-forming bacterium that is considered as a strict anaerobe. It ferments sugars to the organic acids acetate and butyrate or shifts to formation of the solvents - ethanol, butanol and acetone. In most bacteria the major regulator of iron homeostasis is Fur (ferric uptake regulator). Analysis of the genome of Clostridium acetobutylicum has revealed three genes encoding Fur-like proteins. The amino acid sequece of one of them showed 70% similarity to the Fur protein of the closely related Bacillus subtilis.<br>Thus, to gain insight into the role of Fur and the mechanisms for maintenance of iron homeostasis in this strict anaerobic organism, we determined its transcriptional profile in response to iron limitation and inactivation of fur.
Project description:Combination of butanol-hyperproducing and hypertolerant phenotypes is essential for developing microbial strains suitable for industrial production of bio-butanol, i.e. one of most promising liquid biofuels. Clostridium cellulovorans is among the microbial strains with the highest potential for direct production of n-butanol from lignocellulosic wastes, a process that would significantly reduce the cost of bio-butanol. However, butanol exhibits higher toxicity compared to ethanol and C. cellulovorans tolerance to this solvent is low. In the present investigation, comparative proteomics was used to study the response of C. cellulovorans to butanol challenge and understand the tolerance mechanisms activated in this condition. The most important response concerned modulation of protein biosynthesis, folding and degradation. Coherent with previous studies on other bacteria, several heat shock proteins (involved in protein quality control) were upregulated. Globally, our data indicates that protein biosynthesis is reduced, likely not to overload heat shock proteins. Several additional metabolic adaptations were triggered by butanol exposure such as the upregulation of V- and F-type ATPases (involved in ATP synthesis/generation of proton motive force), enzymes involved in amino acid biosynthesis, proteins involved in cell envelope re-arrangement and a redistribution of carbon flux through fermentative pathways. These analyses eventually suggested several potential gene targets for metabolic engineering strategies aimed at improving butanol tolerance in C. cellulovorans.