Natural products with significant biological activities continuously act as rich sources for drug discovery and development.To harness the potential of these valuable compounds,robust methods need to be developed for ...Natural products with significant biological activities continuously act as rich sources for drug discovery and development.To harness the potential of these valuable compounds,robust methods need to be developed for their rapid and sustainable production.Cell-free biosynthesis of pharmaceutical natural products by in vitro reconstruction of the entire biosynthetic pathways represents one such solution.In this review,we focus on in vitro biosynthesis of two important classes of natural products,polyketides(PKs)and nonribosomal peptides(NRPs).First,we summarize purified enzyme-based systems for the biosynthesis of PKs,NRPs,and PK/NRP hybrids.Then,we introduce the cell-free protein synthesis(CFPS)-based technology for natural product production.With that,we discuss challenges and opportunities of cell-free synthetic biology for in vitro biosynthesis of natural products.展开更多
Cell-free system has emerged as a powerful platform with a wide range of in vitro applications and recently has contributed to express metabolic pathways for biosynthesis.Here we report in vitro construction of a nati...Cell-free system has emerged as a powerful platform with a wide range of in vitro applications and recently has contributed to express metabolic pathways for biosynthesis.Here we report in vitro construction of a native biosynthetic pathway for L-4-nitrotryptophan(L-4-nitro-Trp)synthesis using an Escherichia coli-based cell-free protein synthesis(CFPS)system.Naturally,a nitric oxide(NO)synthase(TxtD)and a cytochrome P450 enzyme(TxtE)are responsible for synthesizing L-4-nitro-Trp,which serves as one substrate for the biosynthesis of a nonribosomal peptide herbicide thaxtomin A.Recombinant coexpression of TxtD and TxtE in a heterologous host like E.coli for L-4-nitro-Trp production has not been achieved so far due to the poor or insoluble expression of TxtD.Using CFPS,TxtD and TxtE were successfully expressed in vitro,enabling the formation of L-4-nitro-Trp.After optimization,the cell-free system was able to synthesize approximately 360μM L-4-nitro-Trp within 16 h.Overall,this work expands the application scope of CFPS for study and synthesis of nitro-containing compounds,which are important building blocks widely used in pharmaceuticals,agrochemicals,and industrial chemicals.展开更多
文摘Natural products with significant biological activities continuously act as rich sources for drug discovery and development.To harness the potential of these valuable compounds,robust methods need to be developed for their rapid and sustainable production.Cell-free biosynthesis of pharmaceutical natural products by in vitro reconstruction of the entire biosynthetic pathways represents one such solution.In this review,we focus on in vitro biosynthesis of two important classes of natural products,polyketides(PKs)and nonribosomal peptides(NRPs).First,we summarize purified enzyme-based systems for the biosynthesis of PKs,NRPs,and PK/NRP hybrids.Then,we introduce the cell-free protein synthesis(CFPS)-based technology for natural product production.With that,we discuss challenges and opportunities of cell-free synthetic biology for in vitro biosynthesis of natural products.
基金This work was supported by the National Natural Science Foundation of China(Nos.31971348 and 32171427)the Natural Science Foundation of Shanghai(No.19ZR1477200)J.L.also acknowledges the starting grant from ShanghaiTech University.
文摘Cell-free system has emerged as a powerful platform with a wide range of in vitro applications and recently has contributed to express metabolic pathways for biosynthesis.Here we report in vitro construction of a native biosynthetic pathway for L-4-nitrotryptophan(L-4-nitro-Trp)synthesis using an Escherichia coli-based cell-free protein synthesis(CFPS)system.Naturally,a nitric oxide(NO)synthase(TxtD)and a cytochrome P450 enzyme(TxtE)are responsible for synthesizing L-4-nitro-Trp,which serves as one substrate for the biosynthesis of a nonribosomal peptide herbicide thaxtomin A.Recombinant coexpression of TxtD and TxtE in a heterologous host like E.coli for L-4-nitro-Trp production has not been achieved so far due to the poor or insoluble expression of TxtD.Using CFPS,TxtD and TxtE were successfully expressed in vitro,enabling the formation of L-4-nitro-Trp.After optimization,the cell-free system was able to synthesize approximately 360μM L-4-nitro-Trp within 16 h.Overall,this work expands the application scope of CFPS for study and synthesis of nitro-containing compounds,which are important building blocks widely used in pharmaceuticals,agrochemicals,and industrial chemicals.
