Biosynthesis Of D-glucaric Acid From Sucrose In Metabolically Engineered E.Coli

D-Glucaric acid is a promising platform compound for the synthesis of other value-added chemicals or commercial chemicals.At present,some chemical methods for synthesizing D-glucose diacetate have been put into production,and microbial research is not mature enough.The effect of monosaccharide fermentation is not obvious,the yield is very low,and it is difficult to separate and purify later.In the mixed sugar fermentation,the production of D-glucose diacetate increased,and it is precisely because of this that the method has gained great attention from researchers.Here,we co-expressed cscB,cscA,cscK,inol,miox,udh,and suhB in E.coli BL21(DE3),which are the key genes of sucrose pathway and metabolites products respectively,functionally constructing an unreported route from sucrose to D-glucaric acid.Further,red recombination and Crispr/Cas9 techniques were used to further deletion five metabolic pathway key genes of chromosomal zwf,pgi,ptsG,uxaC,gudD,overexpression of the gene glk compensates for the lack of phosphorylation of D-glucose and the use of a D-fructose-dependent translation control system for the regulation of the gene pgi and determination of Pgi activity at different stages in order to achieve the goal of regulating carbon flux.The above approach ultimately enables recombinant E.coli to use sucrose as the sole carbon source while obtaining higher yields.During the fermentation process,the yield of D-glucaric acid in M9 medium containing?10g/L sucrose was found to be?1.42g,and when sucrose was used as the substrate,the yield was?0.142g/g.