Transcriptional Regulators Of The GntR Family Of Streptomyces Roseosporus For Daptomycin Biosynthesis

Daptomycin,a secondary metabolite of Streptomyces roseosporus fermentation,is a cyclic lipopeptide antibiotic with a completely new bactericidal mechanism.Improving the yield of daptomycin to meet the drug-resistant strain infection drug use has become a struggling goal of researchers.In recent years,the market of daptomycin in China has increased year by year,but at present,the strain producing daptomycin by fermentation has a low titer,which can not meet people's clinical medication needs.With the application of molecular biology,synthetic biology and other technologies,it has become a new trend to engineer strains on the pathway of product biosynthesis,study and analyze the function of genes related to metabolites,and improve the production of microbial natural products.n this paper,we used Streptomyces roseosporus as an experimental agent to study the transcriptional regulators of the GntR family in the daptomycin biosynthetic pathway,as follows:First,the fermentation process was optimized using Streptomyces roseosporus HF-1801 stored in our laboratory in terms of the addition time of precursor capric acid,single addition of capric acid and inoculum size.The feeding time of capric acid was determined to be 48 h after the start of fermentation with a single feed volume of0.15 % and an inoculum size of 4%,and the optimized daptomycin yield reached a maximum of 226 mg/L.Second,to engineer S.roseosporus at the genetic level,a genetic manipulation system was established: the strain was sensitive to all four commonly used antibiotics including Apramycin,a molecular cloning PCR reaction system with annealing temperature of 62 ?and DMSO addition of 5% gave better results,and a S.roseosporus-Escherichia coli conjugative transfer system was established.Finally,SSIG?RS21680,a transcriptional regulator of the GntR family,was selected to study in order to construct the knockout and overexpression strains,which were cultured in different solid media and found that this gene did not have a large effect on the morphological differentiation of the bacteriophage growth,while it was found to negatively regulate the production of daptomycin by fermentation and Cultivation under the same conditions.