Biological Characteristics Of Organophosphate Degrading Bacteria And Enzymology Properties Analysis Of Organophosphate Degradation Enzyme

Phosphorus is an indispensable element in plant growth and development process, but more exist in the combination of the invalid state of phosphorus in the field. Soil microbes was not only able to invalid state of effective phosphorus and phosphorus into free can also degrade the field residues of pesticides.Separation and utilizing the phosphate-solubilizing bacteria in the soil, it can make full use of difficult soluble phosphorus in the soil, create greater benefits and economic benefits.Three organophosphate degrading bacteria Yj1、Yj2 and Yj3 were isolated and purified from soybean rhizosphere soil in this experiment can both degradation soybean lecithin and dimethoate,and has carried on the appraisal,growing conditions optimization, acid phosphatase, alkaline phosphatase and enzyme activity determination of organophosphate degradation enzyme and the separation and purification of organophosphate degrading enzymes of the three kinds of bacteria.On the one hand, can improve the utilization rate of phosphorus removal in the nature, reducing the residue of organophosphorus pesticides in the environment, at the same time to increase the rate of degradation of organophosphorus provide basic theoretical basis. On the other hand, also can lay the foundation for degradation of organic pesticides and organophosphate degradation bacterium agent production,by research the biological characteristics of organophosphate degrading bacteria and the organophosphate degradation enzyme enzymology properties.Via vitek2 automatic analysis system for appraisal, Yj1 is Serratia marcescens and Serratia marcescens WW4(CP003959.1) of 16 s rDNA similarity of 99%.The orthogonal experiment to optimize the medium, get the optimal growth conditions of the strains is the combination of mannose, peptone and pH = 8.Yj1 strain increment is very low under the condition of two kinds of phosphorus source, but soybean lecithin as the phosphorus source of bacteria growth situation is better than that of dimethoate in 72 h.Acid phosphatase, alkaline phosphatase and organophosphate degradation enzyme activity was significantly higher with dimethoate as the phosphorus source of enzyme activity when soybean lecithin as source of phosphorus, and within 72 h acid phosphatase and alkaline phosphatase activity has been higher than organophosphate degrading enzymes.Ammonium sulfate precipitation combined with cation exchange chromatography purified organophosphate degradation enzyme from Yj1 bacteria successfully, SDS-PAGE results show that the purified protein for a single band.And cation exchange chromatography purification ratio is 5.303 times that of the ammonium sulphate precipitation, ammonium sulphate precipitation for the 1.416 times of crude enzyme.Yj2 identified as Acinetobacter and Acinetobacter genomosp. 13(FJ694759.1)of 16 s rDNA similarity of 99%.The orthogonal experiment to optimize the medium, get the optimal growth conditions of the strains is the combination of glucose,(NH4)2SO2 and pH =8. With dimethoate as the phosphorus source within 72 h of Yj2 strain increment is better than that of soybean lecithin.Soybean lecithin as phosphorus acid phosphatase, alkaline phosphatase activity was higher than when the source with dimethoate as the phosphorus source of enzyme activity, but organophosphate degradation enzyme activity is higher than that of soybean lecithin with dimethoate as the phosphorus source. And acid phosphatase activity has been higher than alkaline phosphatase and organophosphate degrading enzymes in 72 h.Ammonium sulfate precipitation combined with cation exchange chromatography purified organophosphate degradation enzyme from Yj2 bacteria successfully, SDS-PAGE results show that the purified protein for a single band.And cation exchange chromatography purification ratio is 10.44 times that of the ammonium sulphate precipitation, ammonium sulphate precipitation for the 1.327 times of crude enzyme.Yj3 identified as Bacillus and Bacillus sp. B2101(JX266369.1)of 16 s rDNA similarity of 99%.The orthogonal experiment to optimize the medium, get the optimal growth conditions of the strains is the combination of glucose, peptone and pH =9. With soybean lecithin as the phosphorus source within 72 h of Yj3 strain increment is better than that of dimethoate.Soybean lecithin as phosphorus acid phosphatase, alkaline phosphatase activity was higher than when the source with dimethoate as the phosphorus source of enzyme activity, but organophosphate degradation enzyme activity is higher than that of soybean lecithin with dimethoate as the phosphorus source. And acid phosphatase activity has been higher than alkaline phosphatase and organophosphate degrading enzymes in 72 h.Ammonium sulfate precipitation combined with cation exchange chromatography purified organophosphate degradation enzyme from Yj3 bacteria successfully, SDS-PAGE results show that the purified protein for a single band.And cation exchange chromatography purification ratio is 7.969 times that of the ammonium sulphate precipitation, ammonium sulphate precipitation for the 1.527 times of crude enzyme.