Chemostat cultivation and transcriptional analyses of Clostridium acetobutylicum mutants with defects in the acid and acetone biosynthetic pathways

• Published in: Applied microbiology and biotechnology 2014-12, Vol.98 (23), p.9777-9794
• Main Authors: Hönicke, Daniel, Lütke-Eversloh, Tina, Liu, Ziyong, Lehmann, Dörte, Liebl, Wolfgang, Ehrenreich, Armin
• Format: Article
• Language: English
• Subjects: Acetone; Acetone - metabolism; Acids; Analysis; Applied Microbial and Cell Physiology; Bacteria; biochemical pathways; Biodiesel fuels; Biomedical and Life Sciences; Biosynthesis; Biosynthetic Pathways - genetics; Biotechnology; Butanol; Butanols - metabolism; Carboxy-Lyases - deficiency; Carboxylic Acids - metabolism; Clostridium; Clostridium acetobutylicum; Clostridium acetobutylicum - genetics; Clostridium acetobutylicum - growth & development; Cultivation; Deoxyribonucleic acid; DNA; Ethanol; Experiments; Fermentation; Gene Expression Profiling; Gene Knockout Techniques; Genes; Genetic aspects; knockout mutants; Life Sciences; Metabolism; Metabolites; microarray technology; Microbial Genetics and Genomics; Microbiology; Mutagenesis; Mutants; Phosphate Acetyltransferase - deficiency; Physiological aspects; Physiology; Production processes; Solvents; Studies; transcription (genetics); Transcription factors; transcriptomics
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-014-6040-9

Clostridium acetobutylicum is a model organism for the biotechnologically important acetone–butanol–ethanol (ABE) fermentation. With the objective to rationally develop strains with improved butanol production, detailed insights into the physiological and genetic mechanisms of solvent production are required. Therefore, pH-controlled phosphate-limited chemostat cultivation and DNA microarray technology were employed for an in-depth analysis of knockout mutants with defects in the central fermentative metabolism. The set of studied mutants included strains with inactivated phosphotransacetylase (pta), phosphotransbutyrylase (ptb), and acetoacetate decarboxylase (adc) encoding genes, as well as an adc/pta double knockout mutant. A comprehensive physiological characterization of the mutants was performed by continuous cultivation, allowing for a well-defined separation of acidogenic and solventogenic growth, combined with the advantage of the high reproducibility of steady-state conditions. The ptb-negative strain C. acetobutylicum ptb::int(87) exhibited the most striking metabolite profile: Sizable amounts of butanol (29 ± 1.3 mM) were already produced during acidogenic growth. The product patterns of the mutants as well as accompanying transcriptomic data are presented and discussed.