Elucidation of the roles of adhE1 and adhE2 in the primary metabolism of Clostridium acetobutylicum by combining in-frame gene deletion and a quantitative system-scale approach

Minyeong Yoo (1) , Christian Croux (2, 3, 4) , Isabelle Meynial Salles (1) , Philippe Soucaille (2, 3, 4)

1 INSA Toulouse - Institut National des Sciences Appliquées - Toulouse
2 LISBP - Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés
3 UT3 - Université Toulouse III - Paul Sabatier
4 Toulouse INP - Institut National Polytechnique (Toulouse)

Background: Clostridium acetobutylicum possesses two homologous adhE genes, adhE1 and adhE2, which have been proposed to be responsible for butanol production in solventogenic and alcohologenic cultures, respectively. To investigate their contributions in detail, in-frame deletion mutants of each gene were constructed and subjected to quantitative transcriptomic (mRNA molecules/cell) and fluxomic analyses in acidogenic, solventogenic, and alcohologenic chemostat cultures. Results: Under solventogenesis, compared to the control strain, only Delta adhE1 mutant exhibited significant changes showing decreased butanol production and transcriptional expression changes in numerous genes. In particular, adhE2 was over expressed (126-fold); thus, AdhE2 can partially replace AdhE1 for butanol production (more than 30 % of the in vivo butanol flux) under solventogenesis. Under alcohologenesis, only Delta adhE2 mutant exhibited striking changes in gene expression and metabolic fluxes, and butanol production was completely lost. Therefore, it was demonstrated that AdhE2 is essential for butanol production and thus metabolic fluxes were redirected toward butyrate formation. Under acidogenesis, metabolic fluxes were not significantly changed in both mutants except the complete loss of butanol formation in Delta adhE2, but numerous changes in gene expression were observed. Furthermore, most of the significantly up-or down-regulated genes under this condition showed the same pattern of change in both mutants. Conclusions: This quantitative system-scale analysis confirms the proposed roles of AdhE1 and AdhE2 in butanol formation that AdhE1 is the key enzyme under solventogenesis, whereas AdhE2 is the key enzyme for butanol formation under acidogenesis and alcohologenesis. Our study also highlights the metabolic flexibility of C. acetobutylicum to genetic alterations of its primary metabolism.

Mots clés: AdhE; Clostridium acetobutylicum; System-scale analysis

Mots clés: Butanol

Domaines: Biotechnologies

Minyeong Yoo, Christian Croux, Isabelle Meynial Salles, Philippe Soucaille. Elucidation of the roles of adhE1 and adhE2 in the primary metabolism of Clostridium acetobutylicum by combining in-frame gene deletion and a quantitative system-scale approach. Biotechnology for Biofuels, 2016, 9, ⟨10.1186/s13068-016-0507-0⟩. ⟨hal-02638685⟩