Abstract:
Application of chemical pesticides has increased significantly worldwide and has raised
serious concerns about environmental pollutions. One of the encouraging trends to
minimize pesticide risk is production of resistant plants containing toxic proteins against
insect pests. Considering the importance of purification and characterization of digestive
enzymes in the production of resistant plants, in this study an α-glucosidase from the
Naranga aenescens Moore's midgut was purified by ammonium sulfate precipitation, ion
exchange chromatography on DEAE-sepharose, and concentrating through
ultrafiltration. The apparent molecular mass of the enzyme was 48 kDa determined by
SDS-PAGE. The optimum pH and temperature of the enzyme were 6.0 and 45°C,
respectively. The irreversible thermoinactivation of the enzyme showed that it was highly
stable at 35ºC but moderately stable at 40 and 45ºC. Zn2+, Hg2+, Co2+ at 10 and 20 mM,
and Ba+2 only in 20 mM strongly inhibited the α-glucosidase activity. Ba2+ and Ca2+ only
at 10 mM, EDTA and Hg2
2+ only at 20 mM and Mg2+ at 10 and 20 mM significantly
increased the enzyme activity. The Km and Kcat values for the α-glucosidase were 0.54 mM
and 3.62 min-1, respectively, when p-Nitrophenyl-α-D-glucopyranoside (pNαG) was used
as a substrate.
Keywords: Ion exchange chromatography, Enzyme activity, Kinetic parameters,
Thermostability Kinetic parameters, Toxic proteins.
