Analysis On Few Key Enzymes From Verticillium Lecanii CA-14 (a New Strain) In Contribution To Biocontrol Mechanism

Verticillium lecanii is an important biocontrol fungi, which can parasitize many kinds of pests and disease fungi such as aphid, rust fungi, Nematoda and so on. It commony parasitizes these pests and disease fungi by enzymes and machinery pressure. Biocontrol mechanism was analyzed from the aspect that extracellular enzymes secreted from Verticillium lecanii CA-14 to control the growth of insect and plant disease fungi. After Verticillium lecanii CA-14 in the extracellular SMCS liquid medium thermostats shaker(200r/min, +/-25℃) were cultured for 48h, 60h and 5d separately, the liquid medium was centrifuged, and the supernatant was put in the steps including ammonium sulfate fractionation, phenyl-Sepharose Fast Flow hydrophobic chromatography, DEAE-Sepharose Fast Flow anionic the exchange column chromatography, CM-Sepharose Fast Flow cation column chromatography, gel electrophoresis and so on, and then a single band of chitinase, cellulose and protease was appeared. The optimum temperature of chitinase was 40°C, the optimum pH range of it was 4.0 to 5.0, it maintained a certain residual activity after incubated in 70℃for 30min. The effects of metal ions on the activity of chitinase was significant, its activity was increased by Ca²⁺,Mg²⁺, Mg²⁺ was the best. The optimum temperature of protease was 37℃, the optimum pH was 9.0, it maintained a certain residual activity after incubated in 70℃for 10min. The effects of metal ions on the activity of protease was significant, its activity was increased by Ca²⁺,Mg²⁺,EDTA, was inhibited by PMSF. The optimum temperature of cellulase was 60°C, the optimum pH of it was 4.8, it maintained a 80% residual activity after incubated in 70℃for 30min. The effects of metal ions on the activity of cellulase was significant, its activity was increased by Ca²⁺,K⁺,Na⁺,Mg²⁺, Ca²⁺,Mg²⁺ were the best, Fe²⁺,Cu²⁺,Zn²⁺ were the worst. The three enzymes could inhibit the germination of plant disease fungi spores, the effects would be better if chitinase and protease were used together, which would be the best if chitinase and inactivated protease were used together.