Purification and Partial Characterization of Three Forms of α-Glucosidase from the Fruit Fly Drosophila melanogaster

Abstract

Three forms of alpha-glucosidase, I, II, and III, have been purified from the whole body extract of adult flies of Drosophila melanogaster in yields of 2.1, 5.3, and 6.7%, respectively. The purification procedures involved ammonium sulfate fractionation, Con A-Sepharose 4B affinity chromatography, DEAE-Sepharose CL-6B ion exchange chromatography, Sephacryl S-200 gel filtration, and preparative gel electrophoresis. Each purified enzyme showed a single band on polyacrylamide gel on both protein and enzyme activity staining. The molecular weights of alpha-glucosidases I, II, and III were estimated to be 200,000, 56,000, and 76,000, respectively, by gel filtration. SDS gels indicated that alpha-glucosidases II and III were each composed of a single polypeptide chain, whereas alpha-glucosidase I was composed of two identical subunits. Both alpha-glucosidases II and III hydrolyzed sucrose and p-nitrophenyl-alpha-D-glucoside (PNPG), but alpha-glucosidase I hydrolyzed PNPG to a much lesser extent than sucrose. For sucrose the pH optima of alpha-glucosidases I, II, and III were pH 6.0, 5.0, and 6.0 and the Km values were 13.1, 8.9, and 10 mM, respectively. For PNPG the pH optima of alpha-glucosidases II and III were pH 5.5 and 6.5 and the Km values were 0.77 and 0.21 mM, respectively.

Neurobiology of the gustatory systems ofDrosophila and some terrestrial insects

“…In Drosophila, a combination of genetic, physiological, and behavioral methods have been used to study chemosensory systems (Benzer, 1973;Falk, 1979;Falk and Atidia, 1975;Quinn et al, 1974;Rodrigues, 1981;Rodrigues andSiddiqi, 1978, 1981;Siddiqi and Rodrigues, 1980;Tanimura and Shimada, 1981;Tanimura et al, 1979;Tompkins, 1979;Tompkins et al, 1979). A large number of mutations affecting the perception of taste have been isolated and characterized behaviorally and genetically (Falk and Atidia, 1975;Isono and Kikuchi, 1974;Morea, 1985;Rodrigues and Siddiqi, 1978;Tanimura et al, 1982;Tompkins et al, 1979).…”

Structural and phylogenetic analysis of α-glucosidase protein in insects

“…This enzyme can be found in the alimentary canal, salivary secretions of insects and hypopharyngeal glands of some insects, such as Apis mellifera (Baker & Lehner 1972;Terra et al 1996). So far, αglus have been isolated and characterized from many insects including Dysdercus peruvianus Guerin-meneville (Hemiptera: Pyrrhocoridae), Sitophilus zeamais Motschulsky (Coleoptera: Curculionidae), A. mellifera L. (Hymenoptera: Apidae), Drosophila melanogaster Meigen (Diptera: Drosophilidae), Glyphodes pyloalis Guenée (Lepidoptera: Pyralidae), Eurygaster integriceps Putton (Hemiptera: Scutelleridae) and Rhopalosiphum padi L. (Homoptera: Aphididae) (Huber & Mathison 1976;Tanimura et al 1979;Baker 1991;Silva & Terra 1997;Ghadamyari et al 2010;Seddigh et al 2012). In the digestive system of Xanthogaleruca luteola, the optimum activity of αglu was reported at pH 5 and temperature 60 • C (Sharifi et al 2011).…”

Multiple receptor proteins for sweet taste inDrosophila discriminated by papain treatment

“…Recently attention has been focused on the primary process of taste, directing the search to the identification of the sugar receptor molecule (Hansen, 1969;Kijima et al, 1977;Tanimura et al, 1979). In the flesh fly, Boettcherisca peregrina, multiple receptor sites on a single sugar receptor cell have been documented (Shimada etal., 1974;Shimada, 1975b;Shimada and Isono, 1978).…”