title: Tunicamycin inhibition of N-glycosylation of α-glucosidase from Aspergillus niveus: partial influence on biochemical properties.
abstract: Treatment of Aspergillus niveus with 30 μg tunicamycin/ml did not interfere with α-glucosidase production, secretion, or its catalytic properties. Fully- and under-glycosylated forms of the enzyme had similar molecular masses, ~56 kDa. Moreover, the absence of N-glycans did not affect either pH optimum (6.0) or temperature optimum (65°C). The K(m) and V(max) values of under- and fully-glycosylated forms of α-glucosidase were similar when assessed for hydrolysis of starch (~0.6 mg/ml, ~350 μmol glucose per min per ml), maltose (~0.54 μmol, ~330 μmol glucose per min per ml) and p-nitrophenyl-α-D: -glucopyranoside (~0.54 μmol, ~8.28 μmol p-nitrophenol per min per ml). However, the under-glycosylated form was sensitive to high temperatures probably because, in addition to stabilizing the protein conformation, glycosylation may also prevent unfolded or partially folded proteins from aggregating. Binding assays clearly showed that the under-glycosylated protein did not bind to concanavalin A but has conserve its jacalin-binding property, suggesting that only O-glycans might be intact on the tunicamycin treated form of the enzyme.
authors.affiliation: Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Avenida Bandeirantes 3900, Monte Alegre, 14040-901, Ribeirão Preto, SP, Brazil.
authors.forename: Tony Márcio
authors.initials: TM
authors.lastname: Silva
authors.forename: Fausto Bruno Dos Reis
authors.initials: FB
authors.lastname: Almeida
authors.forename: André Ricardo de Lima
authors.initials: AR
authors.lastname: Damásio
authors.forename: Alexandre
authors.initials: A
authors.lastname: Maller
authors.forename: Michele
authors.initials: M
authors.lastname: Michelin
authors.forename: João Atílio
authors.initials: JA
authors.lastname: Jorge
authors.forename: Ebert Seixas
authors.initials: ES
authors.lastname: Hanna
authors.forename: Maria Cristina
authors.initials: MC
authors.lastname: Roque-Barreira
authors.forename: Héctor F
authors.initials: HF
authors.lastname: Terenzi
authors.forename: Maria de Lourdes Teixeira de Moraes
authors.initials: Mde L
authors.lastname: Polizeli
mesh_terms: D001230:Aspergillus / Q000187:drug effects* / Q000201:enzymology*; D024202:Electrophoretic Mobility Shift Assay; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D065089:Glycoside Hydrolase Inhibitors; D006031:Glycosylation / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008320:Maltose / Q000378:metabolism; D008970:Molecular Weight; D009598:Nitrophenylgalactosides / Q000378:metabolism; D013213:Starch / Q000378:metabolism; D013696:Temperature; D014415:Tunicamycin / Q000494:pharmacology*; D000520:alpha-Glucosidases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D065089:Glycoside Hydrolase Inhibitors; D014415:Tunicamycin; D009598:Nitrophenylgalactosides; C059200:4-nitrophenylgalactoside; D008320:Maltose; D013213:Starch; D000520:alpha-Glucosidases