Characterization of protoxin protein and chitinase enzyme from indigenous isolates Bacillus thuringiensis

Abstract

Bacillus thuringiensis secreted exochitinase activity when grown in a medium containing chitin. Chitinase and protoxin protein play a important role in the pathogenicity of Bacillus thuringiensis to insect pest. This research was aimed to determined characterization of B. thuringiensis subsp. pakistani indigenous isolates, B. thuringiensis Lot II, and B. thuringiensis 47 to produce chitinase enzyme and protoxin protein. Optimum production of chitinase from B. thuringiensis subsp. pakistani was obtained at 24th hours, pH 7.5, and 40oC. B. thuringiensis Lot II was optimum at 21th hour, pH 6,0, and 50oC. B. thuringiensis 47 was optimum at 21st hour, pH 5,0, and 35 oC. Ammonium sulphate was used for precipitation of chitinase protein. The highest spesific activity of chitinase was obtained on ammonium sulphate at saturation 20% for B. thuringiensis subsp. pakistani, 30% for B. thuringiensis Lot II and 40% for B. thuringiensis 47. The highest production of protoxin protein from B. thuringiensis subsp. pakistani and B. thuringiensis 47 was obtained at 36th hour, whereas B. thuringiensis Lot II at 33rd hour. The molecule weight of chitinase molecule was determined by Sodium Dodecyl Sulfate Solution Polyacrilamide Gel Electrophoresis (SDS-PAGE). Molecular weight of chitinase of B. thuringiensis subsp. pakistani, B. thuringiensis Lot II and 47 estimated at 71,15 kDa. The weight of protoxin protein molecule all isolates estimated 129,99 and 130,04 kDa.