Study On Mutation And Enzymatic Of A Fibrinnolytic Enzyme-Prcducing Strain

A novel plasmin-producing bacterial strain PY,which had been identified as Bacillus LicheniformaoUsing a dosage of about 1 X 1014 to 4X 1015 N ions/cm2 -s and 20kev, the strain B L.PY. was treated by N ion implantation and a stable mutant strain with high yield of plasmin is obtained .In general, the rate of mutation and death grows as the increasing of ion dosage. The shape of strains changed. Shaking fermentation indicates that compared with that of normal strain about 120 strians, the enzymetic production of the mutant one increases 51.13%, reaching 1210U. Fermentation conditions for enzyme production have been established. Composition of the medium (%) is edible sucrose 3, soybean poder 1.2, yeast extract 0.2, MgSO4 7H2O 0.1, CaCl2 0.03, K2HPO4 0.3, KH2PO40.04, pH 7.2, After fermentation in 1.5 liter of fermentorat 37"C with an aeration rate of 1: 0.5 - 1:1 for 4 days, the average fibrinolytic activity was about 1320U/ml culture broth,increase by 65% over the mutant.The purification of enzyme BK include saline extraction,, ion-exchange chromatography on CM-Sephadex C-25 and DEAE-Sephadex A-50, and gel-filtration on Sephadex G-75 ,with a molecular weight of 23 KD. The protocols used were phenols.sulfuric acid method,3,5-dinitrosalicylic acid method and thin layer chromatography of salic gel G. Reducing sugar ang the tatol sugar weremeasured with 3,5-2nitro-salcylic acid.We found that their contents were 7.45% and 7.49% respectnely.The categories of sugar were obtained using silica gel sheet chromatography .The glyco proteins that this enzyme contains mortly were hetrosamine .In the same time,a research was performed to find out how the additionnal metal hydronium would affect the activity of the fibrinolytic enzyme. We could draw an inclusion that EDTA did not have an obvipns inhibitoryaction.These showed that serine was one of the conponents of the active center while metal hydromium wasn't Urea and denaturant SDS inhibited the activity of the enzyme to a degree,but we could still find that the enzyme's ability of anti-denaturant was strong.