Study On Kinds And Characteristics Of Extracellular Protease From Ascosphaera Apis

Bee chalkbrood disease, caused by Ascosphaera apis, was a fungal epidemic disease of honeybee larva. The successful infection of bee larvae depends on the secretion of a variety of extracellular protease. Here we study on the kinds and characteristics of extracellular protease from A. apis strain, ND-2, which was isolated and preservated by Honeybee Physiological and Pathological laboratory, Bee Science College of Fujian Agriculture and Forestry University. Colloid chitin was used to induce the secretion of extracellular protease from Ascosphaera apis, the induced product was precipated by 85% (NH₄)₂SO₄ overnight and then centrifugated 10000 rpm at 4℃for 30 min. The precipitation was dialyzed against double distill water over 24 hours to remove the salt ions and the ions removed product was used as the rude extracellular protease. The induced extracellular protease was certificated containing casein protease, azocasein protease and collagenase. Further, the properties of extracellular protease were studied. The resulted showed that the optimum reaction temperature of casein protease, azocasein protease, collagenase were 45℃,50℃,50℃respectively; the optimum reaction pH were 7,6,8 respectively; Mn²⁺enhanced the activity of casein protease, while the other ions inhibited enzyme activity. Mn²⁺,Zn²⁺,Ca²⁺,Fe²⁺,Mg²⁺ inhibited azocasein protease activity in some degree, while Zn²⁺,Fe²⁺ inhibited significantly. Mn²⁺,Al3+enhanced the collagenase activity, while Zn²⁺,Ca²⁺,Fe²⁺,Mg²⁺ inhibited its activity, among them, Zn²⁺ inhibited significantly.