Reconstruction And Validation For Genome-scale Metabolic Network Of Saccharopolyspora Spinosa

Novel biological pesticide spinosad is a class of macrolide compounds, which issynthesized by Secondary metabolic pathway of Saccharopolyspora spinosa. It hasbecome a global research focus with its unique mechanism and good performance.However, the yield of it is not ideal and can't reach the requirement ofindustrialization due to the limitation of S.spinosa itself, which makes thepop?Larization and application impossible. At present, there is little progress on thegene modification of S.spinosa because of the lack of knowledge on the completemetabolic mechanism of the cell. Genome-scale-metabolic-model, a perfect tool tostudy the metabolic mechanism of the cell, provides an ideal way to investigate thecell metabolism in the whole level. But we have not got theGenome-scale-metabolic-model of S.spinosa?mainly because of the complication ofits metabolism and the lack of genomic information. In this paper, we refered to thegenome annotation information of S.spinosa reported recently and related literaturesto build a Genome-scale-metabolic-model of S.spinosa based on the general modelconstruction method. In this model, it contained1577reactions,1736kinds ofmetabolites,733kinds of enzymes and10major subsystems based on the division ofmetabolic pathway.We use the construted model to conduct a series of caculations, including: theeffect of substrate absorption rate on the spinosad product rate, predicting8aminoacids which can improve the spinosad production and7of them can get experimentdata surporting.In addition, as cofactor NADPH/NADH plays an important part in the regulatorysystem of the synthesis of spinosad. In order to investigate the concret mechanism ofit, we have some special caculations about cofactor NADPH/NADH and NAD(P)+transhydrogenase. The result showed R00112had a great effection on the synthesis ofproducts. In details, the synthesis of spinosad could be promoted by the higheractivity of NAD(P)+-transhydrogenase. According to the calculation result, we overexpressed the genes for NAD(P)+-transhydrogenase in the S.spinosa and then addedthe substrate rhamnose. The fermentation production was increased as expected, which gived a verification of the reliability of the model.