Combinatorial metabolic engineering enables high yield production of α-arbutin from sucrose by biocatalysis

Authors：Zhou, Qi; Wu, Yaokang; Deng, Jieying; Liu, Yanfeng; Li, Jianghua; Du, Guocheng; Lv, Xueqin^*; Liu, Long

Source：Applied Microbiology and Biotechnology, 2023, 107(9): 2897-2910.

DOI：10.1007/s00253-023-12496-2

• alpha-Arbutin
• Bacillus subtilis
• Whole-cell biocatalysis
• PfkA degradation system
• Sucrose conversion

alpha-Arbutin has been widely used as a skin-whitening ingredient. Previously, we successfully produced a-arbutin via wholecell biocatalysis and found that the conversion rate of sucrose to alpha-arbutin was low (similar to 45%). To overcome this issue, herein, we knocked out the genes of enzymes related to the sucrose hydrolysis, including sacB, sacC, levB, and sacA. The sucrose consumption was reduced by 17.4% in 24 h, and the sucrose conversion rate was increased to 51.5%. Furthermore, we developed an inducible protein degradation system with Lon protease isolated from Mesoplasma florum (MfLon) and proteolytic tag to control the PfkA activity, so that more fructose-6-phosphate (F6P) can be converted into glucose-1-phosphate (Glc1P) for a-arbutin synthesis, which can reduce the addition of sucrose and increase the sucrose conversion efficiency. Finally, the pathway of F6P to Glc1P was enhanced by integrating another copy of glucose 6-phosphate isomerase (Pgi) and phosphoglucomutase (PgcA); a high a-arbutin titer (similar to 120 g/L) was obtained. The sucrose conversion rate was increased to 60.4% (mol/mol). In this study, the substrate utilization rate was boosted due to the attenuation of its hydrolysis and the assistance of the intracellular enzymes that converted the side product back into the substrate for a-arbutin synthesis. This strategy provides a new idea for the whole-cell biocatalytic synthesis of other products using sucrose as substrate, especially valuable glycosides.

• Institution
  test; TEST; 江南大学