A Biological Evaluation Of Ni-Trostyrene,Nitrogenous Heterocycle,Phenylhydrazone Compounds Against Magnaporthe Grisea

Rice production has seriously affected by rice blast disease,which is caused by the fungus Magnaporthe grisea.The research about fungicides used to prevent rice blast disease has attracted world-wide attention.Trihydroxynaphthalene reductase(3HNR),which is related to the biosynthesis of vermelone,is the key enzyme in the melanin biosynthetic pathway and has been regarded as an important target enzyme of melanin biosynthesis inhibitors(MBIs).MBIs inhibit the biosynthesis of melanin to prevent the fungus from infecting other plants,and is non-direct antifungal inhibitors.MBIs,such as tricyclazole,which are non-direct fungi-toxic inhibitors,have low levels of resistance with a long service life.Sterol 14? demethylase(CYP51)plays an important role in sterol biosynthesis of eukaryote.It is an important target of rice blast fungicide.Sterol 14a demethylase inhibitors(DMIs)can hinder the biosynthesis of sterol from sterol precursors,so that the structures and functions of the cell membrane can be seriously destroyed.As direct antifungal inhibitors,DMIs,such as azoles,having high bactericidal efficiency and high levels of resistance.In order to combine the advantages of the two kinds of inhibitors,we need to find a novel kind of inhibitor,which can restrain the growth of fungus and inhibit the biosynthesis of melanin at the same time.We aim to find a kind of antifungal compound acting on CYP51 and 3HNR at the same time.The following work has been carried out to fulfill the thesis:1.After heterogeneous expression of 14a-sterol demethylase(CYP51)from Leish-mania infantum,the enzyme was purified by nickel affinity chromatography.The purified protein was analyzed by SDS-PAGE electrophoresis.2.Constructed the system to determine the inhibitor binding spectrum.Biological tests has been carried out on the two enzyme targets named 3HNR and CYP51.According to the results,compound C3S1 has a strong interaction with CYP51 with lowest dissociation constant(Kd),the value is 0.25±0.028?M.Among the inhibitors acting on 3HNR,compound hs32br has the lowest half maximal inhibitory concentration(IC50),the value is 0.29±0.019?M.3.bacteriostasis experiments have been carried out,compound hs65 has the best effect with the lowest EC50,the value is 9.47±0.42ppm.After comprehensive analysis of the results on enzyme level and thalli level,we get experience to optimize the lead compounds in the future.