Important Role of Genome Editing

ABE production globally has experienced different challenges and successes in the past years. Many production plants and industries were established to produce butanol, acetone and ethanol. However, due to the increasing price trend of crude oil, these companies have been shut down in the past four years due to the rapid drop of crude oil price.

Novel and new expertise for cell and process engineering will fast-track the industrial growth of biobutanol production in the future. Solventogenic clostridia are well-studied strains due to their specific ability in alcohols (butanol and ethanol) and acetone biosynthesis. Clostridium acetobutylicum used in ABE fermentation triggered lots of research interests in recent years. Despite the fact that immense efforts have been made to increase butanol yield in the during ABE fermentation, this process still suffers from low butanol yield. 

In order to improve biobutanol production, the butanol-producing abilities of the strains and the corresponding downstream process engineering are key factors (Wang et al., 2014). Though several researchers have worked extensively on this work, there are still limited number of genes that enhance carbon metabolism and sporulation in microbes. Several methods including single cross-over recombination and group II intron retargeting methods have been used. (Sillers et al., 2008; Tummala et al., 2003; Heap et al., 2007). Furthermore, the efficiency of metabolic engineered strains is not acceptable, even not as good as that achieved by the famous mutants Clostridium acetobutylicum JB200 from long-term adjustment and Clostridium beijerinckii BA101 from chemical mutagenesis (Annous and Blaschek, 1991; Xue et al., 2012).

These proofs, presenting less efficiency of rational metabolic engineering compared to traditional random mutagenesis and screening, suggested disadvantaged strain development due to inadequate genetic tools and comprehension of molecular mechanism. With the improvement of genome editing technology and system biology, strain development for ABE fermentation is expected to make a significant breakthrough in coming years. In addition, in situ product recovery techniques lowers the effect of butanol toxicity and efficiently reduce the production cost via continuous removal butanol during fermentation, which will also promote the development of biobutanol production. Recent synthetic biology efforts have successfully introduced butanol toxicity tolerant and butanol- producing genes into various non-native producing microorganisms including E. coli, cyanobacteria, S. cerevisiae, C. tyrobutyricum etc. (Lan and Liao, 2012a, 2012b; Krivoruchko et al., 2013; Yu et al., 2011)

Although these studies have demonstrated the potential and feasibility of applying these not-native hosts for butanol production, the butanol titers and productivities achieved are very low, some of which are disappointingly at least one order of magnitude lower than that achieved by the native butanol-producing strains Clostridium acetobutylicum JB200 or Clostridium beijerinckii BA101. Furthermore, C. acetobutylicum is a typical strain for butanol production, and thus, the genetic manipulation and process engineering strategies developed based on C. acetobutylicum can also be implemented in other species of Clostridium and non-native producing strains. Here, we review the problems and advances in butanol production by C. acetobutylicum, including mechanism and regulation of sugar uptake, metabolic engineering and genome editing for strain improvement, and integrated recovery technologies including conventional single integrated techniques and advanced hybrid recovery strategies.

Based on the summarized work of the past decades, this review highlighted the important role of genome editing and impercipient metabolic regulation in C. acetobutylicum, which could contribute a lot for strain development. In addition, integration of fermentation and recovery aiming to improve the efficiency of biobutanol production was discussed.Wehope this reviewcould facilitate the development and expansion of strain and process engineering formicrobial butanol production, and the final revival of biobutanol production in the future.

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