Production Of Eugenol In Engineered Escherichia Coli

Eugenol is a highly effective plant-derived biological pesticide,which is mainly used to control important agricultural diseases such as tomato gray mold,grape downy mildew and potato late blight.In this paper,ferulic acid-engineered E.coli was used as the chassis cell.Using synthetic biology strategy and modular engineering,the pathway of eugenol was divided into coniferyl alcohol synthesis module and eugenol synthesis module,to conduct gene screening and expression optimization research.We cross-combined three coniferyl acyltransferase genes and four eugenol synthase genes from different plant sources to construct 12 eugenol synthesis modules.Expressed in E.coli BL21(DE3),the optimal module Ph CFAT / Gdl EGS2 was selected.After co-expressed with coniferyl alcohol synthesis module in the ferulic acidengineered E.coli,it was fermented with glucose and de novo synthesis of 44.83 mg/L eugenol.Further,the untranslated regions(UTRs)of the two genes of the eugenol synthesis module were designed.After combination optimization,100 mg/L coniferyl alcohol could be converted into 61.88 mg/L eugenol.Next,we optimized the three genes At4CL1,At CCR1 and yah K of the coniferyl alcohol synthesis module,and expressed them after the monocistronic T7 promoter,and ferulic acid was added as the substrate to synthesize 99.14 mg/L coniferyl alcohol.The optimized eugenol synthesis pathway was expressed in ferulic acid-engineered E.coli,and the initial glucose,yeast extract and inducer concentration were optimized.Under the appropriate medium and fermentation conditions,the yield of eugenol was 103.64mg/L.At present,the preparation of eugenol from clove and other plants in China has the problems of slow growth and other raw materials.This article combines metabolic engineering and synthetic biology to realize the de novo synthesis of eugenol by microorganisms,which has important application value and industrialization prospects.