Heterologous Pathways Design Based On Integrated Network Model

Design of heterologous synthetic pathways for high value-added products to expand the metabolic potential of host organisms is an important strategy for metabolic engineering and synthetic biology.Traditional metabolic engineering is an inefficient process for pathway design and strain construction because it is only based on biochemical knowledge and trial and error.In order to solve this problem,this study selected the high-quality genome-scale metabolic network model from Bi GG database as the chassis strain models,and then constructed the integrated network model BKMD by integrating and modifying reactions from Bi GG,KEGG and Meta Cyc databases.Based on a chassis model and the integrated network model,a weight-based parsimonious flux balance analysis(wp FBA)algorithm was developed,and a web server WPHP(https://wphp.biodesign.ac.cn)was built for heterologous pathway design.Firstly,the high quality integrated network model BKMD and chassis model library were constructed.Reactions information in Bi GG,KEGG and Meta Cyc databases was collected,simplified and modified to ensure the accuracy of pathway design.The mapping relationships between reactions and metabolites in Bi GG,Meta Cyc,Meta Net X and RAVEN were collected and integrated.With the help of the mapping relationships,the redundant reactions deletion and metabolite ID conversion in the process of databases integration are completed.Then a high-quality integrated network model BKMD containing 64749 reactions and 36058 metabolites was constructed,extending the scope of pathway design.In addition,109 high-quality genome-scale metabolic network models were collected,and the medium components and respiratory chains of the models were normalized to construct the chassis models database.Then we developed the wp FBA algorithm.By increasing the weight of the reactions in the Bi GG-derived chassis models and reducing the weight of reactions in the integrated network model BKMD,the algorithm can design the minimal amount of heterologous reactions required for the synthesis of heterologous products based on specific chassis cells,and design multiple feasible synthesis pathways for products.Finally,an online pathways design website WPHP was built based on the integrated network model BKMD,chassis model library and wp FBA algorithm.The synthetic pathways of products can be designed by setting the chassis model,substrate,product and maximum number of heterologous reactions.Using E.coli,P.putida,C.glutamate and S.cerevisiae as the chassis,supplying glucose,glycerol and xylose respectively,the pathways of 280 products were designed and the pathways of ginsenoside CK,taxifolin and itaconic acid were analyzed in detail.By comparing the published methods and analyzing the results,it was found that WPHP has advantages in guiding the selection of chassis strains,exploring new pathways,guiding the utilization of substrates and discovering the minimal heterologous reactions required for the synthesis of heterologous products.