Molecular cloning and characterization of the α-glucosidase II from Bombyx mori and Spodoptera frugiperda

The alpha-glucosidase II (GII) is a heterodimer of alpha- and beta-subunits and important for N-glycosylation processing and quality control of nascent glycoproteins. Although high concentration of alpha-glucosidase inhibitors from mulberry leaves accumulate in silkworms (Bombyx mori) by feeding, silkworm does not show any toxic symptom against these inhibitors and N-glycosylation of recombinant proteins is not affected. We, therefore, hypothesized that silkworm GII is not sensitive to the alpha-glucosidase inhibitors from mulberry leaves. However, the genes for B. mori GII subunits have not yet been identified, and the protein has not been characterized. Therefore, we isolated the B. mori GII alpha- and beta-subunit genes and the GII alpha-subunit gene of Spodoptera frugiperda, which does not feed on mulberry leaves. We used a baculovirus expression system to produce the recombinant GII subunits and identified their enzyme characteristics. The recombinant GII alpha-subunits of B. mori and S. frugiperda hydrolyzed p-nitrophenyl alpha-D-glucopyranoside (pNP-alpha Glc) but were inactive toward N-glycan. Although the B. mori GII beta-subunit was not required for the hydrolysis of pNP-alpha Glc, a B. mori GII complex of the alpha- and beta-subunits was required for N-glycan cleavage. As hypothesized, the B. mori GII alpha-subunit protein was less sensitive to alpha-glucosidase inhibitors than was the S. frugiperda GII alpha-subunit protein. Our observations suggest that the low sensitivity of GII contributes to the ability of B. mori to evade the toxic effect of alpha-glucosidase inhibitors from mulberry leaves. (C) 2013 Elsevier Ltd. All-rights reserved.