Reconstruction Of Saccharopolyspora Erythraea Genome-scale Metabolic Model And Its Application

Erythromycin is an important broad-spectrum 14-membered macrolide antibiotic which has been widely used in the treatment of many diseases caused by pathogenic Gram-positive bacteria.Saccharopolyspora erythraea?S.erythraea?,formerly called Streptomyces erythraeus,is a kind of Gram-positive filamentous bacterium which can produce the medical useful antibiotic erythromycin.Genome-scale metabolic models?GEMs?has become an indispensable tool for cellular metabolic network research in systems biology.It is helpful to understand the metabolic regulation and improve the existing processes of S.erythraea.Furthermore,the local approach can be used to improve the production of erythromycin through regulation of the gene metabolic targets in S.erythraea.GEMs model can also be used to predict the utilization of metabolic substrates of S.erythraea,further develop a clear and controllable chemically dedined medium for industrial development.Due to the reason that there are many problems such as difficulty in operation,incomplete in structure,low profiles in model coverage and no data validation of wet experimental phenotype,the newly constructed model iZZ1342 has made many aspects of upgrading,and a series of convincing validations have been carried out to ensure the high quality of the reconstruction GEMs model of S.erythraea.Firstly,the latest and whole genome annotation information of S.erythraea from different databases?KEGG,NCBI,Enzyme and UniprotKB?were extracted.Moreover,the annotated genes which are lacked in the original version of model are added,and the manually refinement on the preliminarily constructed model was carried out.The final parameters of iZZ 1342,which was constructed in the S.erythraea GEMs model,were summarized as follows:1342 annotated genes,1551 metabolic reactions and 1614 metabolites.Compared with the original model,the number of total effective reactions and metabolites are significantly increased.Next,for the newly constructed model iZZ1342,five aspects of verification have been carried out in this reasearch.Compared with the transcriptomics data obtained from the published literature,86.3%ORFs and 92.9%reactions in iZZ1342 can be verified.The results of the sensitivity analysis showed the similar trend in the E.coli GEMs.The prediction of growth on available 27 kinds of carbon sources and 33 kinds of nitrogen sources showed the accuracy rate was 77.8%and 87.9%respectively.Compared with the physiological data obtained from chemostat cultivation,the simulation results showed good consistency.The correlation coefficient between the 13C-labeled experiment data and the flux simulation result was 0.97.All the above results show that the iZZ1342 model has good performance.Thirdly,the reconstructed model iZZ1342 was used to predict the essential genes and the prediction results was used to find the key gene targets in the specific metabolic pathways.Through the computer aided simulation,89 semi-essential genes ware found in the intracellular metabolic pathways of S.erythraea.In order to validate the effect on the synthesis of erythromycin after knocking out these gene targets,this research selected some metabolic genes targets which have been knocked out in our laboratory?the experiments have been completed?or the published literature to verify the accuracy of the prediction.These targets include:SACE₅639,SACE₀728,SACE₀731 and SACE₆669.Combined the model data with the data in the literature,the results showed that the calculation results of three metabolic targets,i.e.SACE₅639,SACE₀731 and SACE₆669 are consistent well with those reported in the literature.However,the calculation result of SACE₀728 metabolic target is inconsistent with the results in the literature,and further analyses the reasons for the non-conformity may be related to the complex regulation of secondary metabolism.The results of prediction showed that the iZZ1342 model can be used for the prediction of key gene targets,and the accuracy is up to a high level.The above results showed that the iZZ1342 model has good performance and lays a good foundation for the biological research of S.erythraea system.