Breeding Mutants Of Streptomyces Fungicidicus By Genome Shuffling For Enduracidin Production

Enduracidin,produced by soil bacterium Streptomyces fungicidicus No.B5477.is one kind of lipodepsipeptides and composed by a 17 amino acids conjunted with fatty acids.The development of the product has the good economic and social performance for its high safety,toxicity,low-residue and active to a wide variety of Gram-positive bacteria.Genome shuffling was conducted to improve the producers with high productivity of enduracidin.The shuffled strain S.fungicidious EL-30 was obtained and promised a evaluated yield.And specially,influences of metal ions on enduracidin synthesis and hyphea pellet performance were investigated.After five rounds of protoplast fusion,five high-yielding strains were obtained,of which fusant EL-10,EL-12,EL-18,EL-22,and EL-30 produced enduracidin up to 1950.8,1796.4,1834.9,1234.1 and 1954.5 mg/L respectively.Enduracidin yields were of 4.62,4.23,4.35,2.92 and 4.63 times to that of the original strain L-35.The enduracidin productivity of the strain is stable after five passage strains.It was suggested that these genome shuffled strains would be indudtrial potential.Fermentation with the shuffled strain S.fungicidious EL-30 was performed in 3.7 liter fermentor,the yield of enduracidin reached 2.66 g/L.The effects of different metal ions on cell growth and enduracidin production of S.fungicidious EL-30 were studied.The Mn2+ was chose for further research for its positive influence on enduracidin biosynthesis.Then the effects of Mn2+ on the organic acids and amino acids catabolism during the enduracidin biosynthesis were investigated.The results showed that concentrations of organic acids in fermentation with 2 mmol/L Mn2+ supplement changed greatly.To the detected organic acids their concentrations were improved.They are utilized as precursors for amino acid biosynthesis,and then fufil into the precursor pool for enduracidin.Furthermore,the influence of Mn2+ on mycelium differentiation in submerged cultures of S.fungicidious EL-30 was investigated.The results showed that omission of Mn2+ from the fermentation medium resulted in dense mycelium pellet with large diameter,and most of the periphery of the pellet cracked into fragments at 144 h to 156 h.In the presence of Mn2+(2 mmol/L),mycelium pellet was loose obviously with small diameter which benefit for the oxygen and nutrient's transferring in the mycelium pellet.At 144 h to 156 h,the periphery of the pellet was still stable and has a longer growth arrest time which beneficial for the enduracidin accumulation.