Background
Acetoin (AC) is a vital platform chemical widely used in food, pharmaceutical and chemical industries. With increasing concern over non-renewable resources and environmental issues, using low-cost biomass for acetoin production by microbial fermentation is undoubtedly a promising strategy.
Results
This work reduces the disadvantages of Bacillus subtilis during fermentation by regulating genes involved in spore formation and autolysis. Then, optimizing intracellular redox homeostasis through Rex protein mitigated the detrimental effects of NADH produced by the glycolytic metabolic pathway on the process of AC production. Subsequently, multiple pathways that compete with AC production are blocked to optimize carbon flux allocation. Finally, the population cell density-induced promoter was used to enhance the AC synthesis pathway. Fermentation was carried out in a 5-L bioreactor using bagasse lignocellulosic hydrolysate, resulting in a final titer of 64.3 g/L, which was 89.5% of the theoretical yield.
Conclusions
The recombinant strain BSMAY-4-PsrfA provides an economical and efficient strategy for large-scale industrial production of acetoin.
α-Arbutin is extensively used in cosmetic industries.
The
lack of highly active enzymes and the cytotoxicity of hydroquinone
limit the biosynthesis of α-arbutin. In this study, a whole-cell
biocatalytic approach based on enzyme engineering and engineered cell
modification was identified as effective in enhancing α-arbutin
production. First, a sucrose phosphorylase (SPase) mutant with higher
enzyme activity was obtained by experimental screening. Next, to avoid
the oxidation of hydroquinone, we established an anaerobic process
to improve the robustness of the cells by knocking out lytC, sdpC, and skfA in Bacillus subtilis and overcoming the inhibitory effect
of a high concentration of hydroquinone. Finally, the engineered strain
was used for biotransformation in a 5 L fermenter with batch feeding
for 24 h. The final yield of α-arbutin achieved was 129.6 g/L,
which may provide a basis for the large-scale industrial production
of α-arbutin.
Bacillus subtilis is a gram-positive bacterium, a promising microorganism due to its strong extracellular protein secretion ability, non-toxic, and relatively mature industrial fermentation technology. However, cell autolysis during fermentation restricts the industrial application of B. subtilis. With the fast advancement of molecular biology and genetic engineering technology, various advanced procedures and gene editing tools have been used to successfully construct autolysis-resistant B. subtilis chassis cells to manufacture various biological products. This paper first analyses the causes of autolysis in B. subtilis from a mechanistic perspective and outlines various strategies to address autolysis in B. subtilis. Finally, potential strategies for solving the autolysis problem of B. subtilis are foreseen.
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