Biosynthesis Of Phenol,tyrosol And Hydroxytyrosol Using The E.coli Shikimate Pathway

Aromatic compounds are kind of organic compounds containing aromatic benzene ring or heterocyclic ring.The metabolic engineering of microbial synthetic aromatic compounds is a promising altermative method because the chemical synthesis of certain aromatic compounds can cause serious environmental pollution and threaten human life.Some stydies have shown that the ubiquitous microbial shikimate pathway can provide precursors for the biosynthesis of some aromatic componds.Therefore,in this paper,the non-natural metabolic biosynthetic pathways for bio-production of the three aromatic compounds of phenol,tyrosol and hydroxytyrosol was designed and constructed using the shikimate pathway in E.coli.The main research results of this study are as follows.1.The recombinant plasmid pRSFDuet-Ubic-bsdB-bsdC-bsdD was constructed in which the gene Ubic from E.coli K12 expressed chorismate pyruvatelyase and the three genes bsdB,bsdC and bsdD coexpressed p-hydroxybenzoate decarboxylase.The de novo synthesis of phenol was successfully achieved with E.coli BL21(DE3)as the host strain and its yield was 14.3 mg/L.Finally,tributyrin was used as the extractant and the yield of phenol in a two-phase extractive fermentation was increase to 45.2%,reaching 26.1 mg/L.2.The recombinant plasmid PETDuct-AROIO was constructed,and the gene AROIO from Saccharomyces cerevisiae S288C expressed.p-hydroxyphenylpyruvate decarboxylase.The de novo synthesis of tyrosol was successfully achieved with E.coli JM109(DE3)as the host strain and its yield was 46.7 mg/L.3.The recombinant plasmid pETduet-ARO10-hpaB-hpaC was constructed,and the gene hpaB and hpaC from Pseudomonas aeruginosa PA01 coexpressed 4-hydroxyphenylacetate-3-hydroxylase.The de novo synthesis of hydroxytrosol was achieved based on the production of tyrosol and with E.coli JM109(DE3)as the host strain and its yield was 4.9 mg/L.4.The gene FeaB was knocked out by using the ?-red recombination system in E.coli JM109(DE3),blocking the formation of by-products.Then,the yields of tyrosol and hydroxytyrosol were increasing by 29.5%and 56.2%,reaching 66.3 mg/L and 11.2 mg/L.After supplementing 1 mM tyrosine in vitro,their yields were increasing by 73.0%and 39.5%,reaching 245.7 mg/L and 18.5 mg/L.