PHENOL BETA-GLUCOSYLTRANSFERASE AND BETA-GLUCOSIDASE ACTIVITIES IN THE TOBACCO HORNWORM LARVA MANDUCA-SEXTA (L) - PROPERTIES AND TISSUE LOCALIZATION

Phenol beta-glucosyltransferase (PGT; EC 2.4.1.35) was studied in feeding fifth stadium larvae of the tobacco hornworm, Manduca sexta, using reversed-phase HPLC with absorbance detection to separate and quantify both the model substrate, p-nitrophenol (PNP), and the product, p-nitrophenyl beta-D-glucopyranoside (PNP-Glc). About 90% of total PGT activity in tissue homogenates was associated with the particulate fraction (15,000g), with the remainder in the microsomal fraction. PGT activity was observed in all tissues, with highest activities in the labial gland and fat body. Appreciable activity occurred in midgut and hindgut tissue but none was found in hemolymph. PGT activity was also observed in eggs and larval fat body at different times during development. Activity was optimal at pH 7.5-9 and was highest with UDP-Glc as a glucose donor. However, appreciable PGT activity was observed with dTDP-Glc or GDP-Glc in place of UDP-Glc. The divalent cations Ca2+, Co2+, Mg2+, and Mn2+ stimulated activity, whereas Zn2+ and Hg2+, as well as pretreatment with the detergent Triton X-100, were inhibitory. Endogenous beta-glucosidase in the PGT-enriched fractions, especially from the midgut, antagonized the beta-glucosylation process, and interference was minimized with higher pH and the addition of D-gluconic acid lactone to the incubation mixtures. The possible role of transglucosylation in the detoxication of phenolic xenobiotics and biotransformation of endogenous phenolic compounds in insects is discussed. Comparisons of PGT with some glycosyltransferases involved in endogenous and xenobiotic conjugation in insects and other organisms are reviewed.