Metabolic engineering of the non-sporulating, non-solventogenic Clostridium acetobutylicum strain M5 to produce butanol without acetone demonstrate the robustness of the acid-formation pathways and the importance of the electron balance

Metabolic Engineering
Ryan SillersEleftherios T Papoutsakis

Abstract

The primary alcohol/aldehyde dehydrogenase (coded by the aad gene) is responsible for butanol formation in Clostridium acetobutylicum. We complemented the non-sporulating, non-solvent-producing C. acetobutylicum M5 strain (which has lost the pSOL1 megaplasmid containing aad and the acetone-formation genes) with aad expressed from the phosphotransbutyrylase promoter and restored butanol production to wild type levels. Because no acetone was produced, no acids (acetate or butyrate) were re-assimilated leading to high butyrate but especially acetate levels. To counter acetate production, we examined thiolase overexpression in order reduce the acetyl-CoA pool and enhance the butyryl-CoA pool. We combined thiolase overexpression with aad overexpression aiming to also enhance butanol formation. While limiting the formation of acetate and ethanol, the butanol titers were not improved. We also generated acetate kinase (AK) and butyrate kinase (BK) knockout (KO) mutants of M5 using a modified protocol to increase the antibiotic-resistance gene expression. These strains exhibited greater than 60% reduction in acetate or butyrate formation, respectively. We complemented the AKKO M5 strain with aad overexpression, but could not successfull...Continue Reading

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Citations

Aug 4, 2009·Journal of Bacteriology·Falk HillmannHubert Bahl
Jun 7, 2011·Applied and Environmental Microbiology·Cong T TrinhDouglas S Clark
Jun 30, 2009·Journal of Industrial Microbiology & Biotechnology·Yan-Ning ZhengDong-Zhi He
Jan 18, 2012·Applied Microbiology and Biotechnology·Wouter KuitGerrit Eggink
May 12, 2012·Applied Microbiology and Biotechnology·Dörte LehmannTina Lütke-Eversloh
Sep 10, 2013·Applied Microbiology and Biotechnology·Yu-Sin JangSang Yup Lee
Sep 18, 2012·Metabolic Engineering·Clare M CooksleyNigel P Minton
Aug 22, 2012·Metabolic Engineering·Antje MayHubert Bahl
May 17, 2011·Metabolic Engineering·Ethan I Lan, James C Liao
May 3, 2011·Metabolic Engineering·Mingrui YuShang-Tian Yang
Mar 8, 2011·Current Opinion in Biotechnology·Tina Lütke-Eversloh, Hubert Bahl
May 26, 2009·Metabolic Engineering·David R NielsenKristala L Jones Prather
Dec 9, 2008·Metabolic Engineering·Gregory Stephanopoulos
Sep 2, 2008·Current Opinion in Biotechnology·Eleftherios T Papoutsakis
Jul 16, 2015·Biotechnology and Bioengineering·Timothy KernanScott Banta
Oct 26, 2012·Biotechnology and Bioengineering·Miriam S Mann, Tina Lütke-Eversloh
Jul 27, 2012·Biotechnology and Bioengineering·Corjan van den BergAdrie J J Straathof
May 18, 2012·TheScientificWorldJournal·Mohammed DwidarByoung-In Sang
Jul 2, 2014·Biotechnology and Bioengineering·András PásztorPatrik R Jones
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Dec 3, 2014·Journal of Microbiological Methods·Ning ZhangSheng Yang
Apr 9, 2015·Applied Microbiology and Biotechnology·Le YuShang-Tian Yang

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