Alleviation of Carbon Catabolite Repression through araR and xylR Inactivation in Clostridium acetobutylicum DSM 792

• Published in: Applied and environmental microbiology Vol. 89; no. 3; p. e0213522
• Main Authors: Delarouzée, Alexandre, Lopes Ferreira, Nicolas, Wasels, François
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 29.03.2023
• Subjects: Arabinose; Assimilation; Bioconversion; Biotechnology; Carbohydrates; Carbon; Catabolite Repression; Chemical Sciences; Clostridium acetobutylicum; Clostridium acetobutylicum - genetics; Deactivation; Deletion; Fermentation; Genes; Genetic modification; Glucose; Growth rate; Inactivation; Lignocellulose; Microorganisms; Pentose; Repressors; Substrates; Sugar; Sugars; Transcription factors; Xylose; Clostridium acetobutylicum; catabolite repression; CRISPR-Cas9; Catabolite Repression; Clostridium Acetobutylicum
• ISSN: 0099-2240; 1098-5336
• DOI: 10.1128/aem.02135-22

Efficient bioconversion processes of lignocellulose-derived carbohydrates into chemicals have received increasing interest in the last decades since they represent a promising alternative to petro-based processes. Despite efforts to adapt microorganisms to the use of such substrates, one of their major limitations remains their inability to consume multiple sugars simultaneously. In particular, the solventogenic model organism Clostridium acetobutylicum struggles to efficiently use second generation (2G) substrates because of carbon catabolite repression mechanisms that prevent the assimilation of xylose and arabinose in the presence of glucose. In this study, we addressed this issue by inactivating genes encoding transcriptional repressors involved in such mechanisms in the C. acetobutylicum strain DSM 792. Our results showed that the deletion of the two putative copies of (CA_C2613 and CA_C3673) had little or no effect on the ability of the strain to consume xylose. Unlikely, the deletion of (CA_C1340) led to a 2.5-fold growth rate increase on xylose. The deletion of both and genes resulted in the coassimilation of arabinose together with glucose, while xylose consumption remained inefficient. Transcriptional analyses of the wild-type strain and mutants grown on glucose, arabinose, xylose, and combinations of them provided a crucial, global overview of regulations triggered by the products of both and in C. acetobutylicum. As suggested by these data, overexpression of and led to further improvement of pentose assimilation. Those results represent a step forward in the development of genetically modified strains of C. acetobutylicum able to coassimilate lignocellulosic-derived sugars. C. acetobutylicum is a strong candidate to produce chemicals of interest such as C3 and C4 alcohols. Used for more than a century for its capacity to produce a mixture of acetone, butanol, and ethanol from first generation (1G) substrates, its natural ability to assimilate a wide variety of monoosides also predisposes it as an auspicious organism for the valorization of lignocellulose-derived sugar mixtures. To achieve this purpose, a better understanding of carbon catabolite repression mechanisms is essential. The work done here provides critical knowledge on how these mechanisms occur during growth on glucose, arabinose, and xylose mixtures, as well as strategies to tackle them.