Heterologous biosynthesis of natural product naringenin by co-culture engineering

Co-culture engineering is an emerging approach for microbial biosynthesis of a variety of biochemicals.In this study,E.coli-E.coli co-cultures were developed for heterologous biosynthesis of the natural product naringenin.The co-cultures were composed of two independent E.coli strains dedicated to functional expression of different portions of the biosynthetic pathway,respectively.The co-culture biosynthesis was optimized by investigating the effect of carbon source,E.coli strain selection,timing of IPTG induction and the inoculation ratio between the co-culture strains.Compared with the monoculture strategy,the utilization of the designed co-cultures significantly improved the naringenin production,largely due to the reduction of metabolic stress,employment of proper hosts for improving pathway enzyme activities,and flexible adjustment of the relative biosynthetic strength between the coculture strains.The findings of this study extend the applicability of co-culture engineering in complex natural product biosynthesis.

Compound Danshen Yeast 1.0

Background:Heterologous synthesis presents a promising new approach for accessing the active ingredients of traditional Chinese medicine(TCM),contributing to the conservation of natural medicinal resources.Compound Danshen preparation,a widely used TCM formulation,is designed to treat coronary heart disease and angina pectoris.It is primarily composed of Salviae Miltiorrhizae Radix et Rhizoma(Danshen),Notoginseng Radix et Rhizoma(Sanqi),and borneol(Bingpian).Danshen primarily yields tanshinones and phenolic acids,while Sanqi produces notoginsenosides.Borneol serves as an auxiliary agent to promote mental clarity,dissipate heat,and relieve pain.Objectives:The objective is to employ heterologous synthesis in a single yeast strain to produce the active ingredients of Compound Danshen preparation,and these include notoginsenosides,tanshinones,and borneol.Methods:Firstly,the“plug-and-play”terpene synthase screening framework in Saccharomyces cerevisiae was used for the identification of a novel,highly efficient bornyl diphosphate synthase.Furthermore,by leveraging the Compound Danshen preparation as a basis to concurrently reconstruct the biosynthetic pathways for the notoginsenoside precursor protopanaxadiol(a triterpene),the tanshinone precursor miltiradiene(a diterpene),and borneol(a monoterpene)within a single yeast strain.Results:This engineered strain,termed Compound Danshen Yeast 1.0,successfully produced protopanaxadiol at 62.34 mg/L,miltiradiene at 15.38 mg/L,and borneol at 2.54 mg/L in shake-flask fermentation.Conclusions:This research signifies the inaugural cross-species andmulticomponent synthetic biology endeavor that enables the synthesis of active ingredients in engineered cells,setting the stage for the industrial manufacture of TCM compounds.