Genome analysis of a hyper acetone‐butanol‐ethanol (ABE) producing Clostridium acetobutylicum BKM19

Abstract

Previously the development of a hyper acetone-butanol-ethanol (ABE) producing Clostridium acetobutylicum BKM19 strain capable of producing 30.5% more total solvent by random mutagenesis of its parental strain PJC4BK, which is a buk mutant C. acetobutylicum ATCC 824 strain is reported. Here, BKM19 and PJC4BK strains are re-sequenced by a high-throughput sequencing technique to understand the mutations responsible for enhanced solvent production. In comparison with the C. acetobutylicum PJC4BK, 13 single nucleotide variants (SNVs), one deletion and one back mutation SNV are identified in the C. acetobutylicum BKM19 genome. Except for one SNV found in the megaplasmid, all mutations are found in the chromosome of BKM19. Among them, a mutation in the thlA gene encoding thiolase is further studied with respect to enzyme activity and butanol production. The mutant thiolase (thlA ) is showed a 32% higher activity than that of the wild-type thiolase (thlA ). In batch fermentation, butanol production is increased by 26% and 23% when the thlA gene is overexpressed in the wild-type C. acetobutylicum ATCC 824 and in its derivative, the thlA-knockdown TKW-A strain, respectively. Based on structural analysis, the mutation in thiolase does not have a direct effect on the regulatory determinant region (RDR). However, the mutation at the 5 residue seems to influence the stability of the RDR, and thus, increases the enzymatic activity and enhances solvent production in the BKM19 strain.

Simultaneous saccharification and fermentation of sago hampas into biobutanol by Clostridium acetobutylicum ATCC 824

“…This observation might be due to the presence of a variety of sugars existed in the sago hydrolysate. Although Clostridia can consume both hexose and pentose sugars for their growth and products formation, these microorganisms show diauxic behavior, by which they consume glucose first followed by xylose . Ezeji et al also reported that Clostridia prefer glucose than C5‐sources when tested in ABE fermentation model.…”

TargeTron Technology Applicable in Solventogenic Clostridia: Revisiting 12 Years’ Advances

“…Table 2 depict the genes disrupted in the ABE metabolic pathway along with the phenotypic changes of the mutants. Pta, ptb, ack, and buk are genes involved in the acetate and butyrate synthesis, which are usually inactivated to reduce acid production and drive more carbon source to ethanol/butanol [15,54,64,[69][70][71][72].…”

Effects of orphan histidine kinases on clostridial sporulation progression and metabolism

“…However, owing to poor understanding of the clostridial physiology and metabolism, industrial bio‐butanol production by solventogenic clostridia is still challenging because its solventogenic ability is limited by sporulation (Yang et al, 2018), which leads to low and unstable butanol production and a high production cost (Zhao et al, 2013). Various genome‐scale models (Dash et al, 2016; Liao et al, 2015; Yoo et al, 2015) and omics analyses (Cho et al, 2017; Hu et al, 2011; Mao et al, 2010; Paredes et al, 2005) focusing on C. acetobutylicum ATCC 824 have been used to shed light on the complex physiology and metabolic and regulatory networks involved in solventogenesis and sporulation, which is the main survival strategy for cells to escape from environmental stress (Amador‐Noguez et al, 2011).…”