Synthetic Biology on Acetogenic Bacteria for Highly Efficient Conversion of C1 Gases to Biochemicals

Abstract

Synthesis gas, which is mainly produced from fossil fuels or biomass gasification, consists of C1 gases such as carbon monoxide, carbon dioxide, and methane as well as hydrogen. Acetogenic bacteria (acetogens) have emerged as an alternative solution to recycle C1 gases by converting them into value-added biochemicals using the Wood-Ljungdahl pathway. Despite the advantage of utilizing acetogens as biocatalysts, it is difficult to develop industrial-scale bioprocesses because of their slow growth rates and low productivities. To solve these problems, conventional approaches to metabolic engineering have been applied; however, there are several limitations owing to the lack of required genetic bioparts for regulating their metabolic pathways. Recently, synthetic biology based on genetic parts, modules, and circuit design has been actively exploited to overcome the limitations in acetogen engineering. This review covers synthetic biology applications to design and build industrial platform acetogens.

Production of the biocommodities butanol and acetone from methanol with fluorescent FAST-tagged proteins using metabolically engineered strains of Eubacterium limosum

“…Molecular tools with different levels of possibilities for metabolic engineering are available for acetogens such as A. woodii , Clostridium ljungdahlii , C. autoethanogenum , and E. limosum. That includes, e.g., genomic editing tools [ 10 – 14 ] and the expression of recombinant pathways to produce biocommodities such as butanol [ 15 , 16 ], acetone [ 17 – 19 ], isopropanol [ 20 ], 3-hydroxybutyrate [ 21 ], or poly(3-hydroxybutyrate) [ 22 ]. However, fluorescent reporter systems which are well-established and often used tools in molecular biology to study gene expression [ 23 , 24 ], promoter activities [ 25 , 26 ], or the dynamics in microbial populations and co-cultures [ 27 – 29 ] are still restricted for acetogens, basically due to the lack of proteins that show bright fluorescence under anaerobic conditions.…”

Isobutanol Production by Autotrophic Acetogenic Bacteria

“…Thus, 2,3butanediol is the prime candidate for a product to be eliminated. If the ClosTron technology does not work (as mentioned, our attempt to inactivate the ilvC gene failed), CRISPR/Cas9-based genome editing will be an alternative that has recently been developed for C. ljungdahlii (Huang et al, 2016;Jin et al, 2020). The optimal temperatures for both key enzymes, ketoisovalerate decarboxylase (45 • C) and ketoisovalerate ferredoxin oxidoreductase (growth temperature of C. thermocellum is 60 • C), significantly exceed the range of mesophilic fermentations.…”

Carbon-negative production of acetone and isopropanol by gas fermentation at industrial pilot scale

“…Since the first demonstration of genetic engineering of an acetogen in 2010 (ref. 28 ), considerable advances in the genetic toolbox of acetogens have been made 29,30 . Genetic engineering of acetogens has enabled small-scale production of more than 50 molecules from C1 gases, including industrially important alcohols (butanol, hexanol, IPA, etc.…”

“…), ketones (acetone and methyl ethyl ketone) and dienes (butylene, isoprene, etc.) 15,[17][18][19][28][29][30] . However, efforts to develop efficient production strains have been hampered by the lack of high-throughput strain engineering workflows and a pathway to industrial scale-up 29 .…”