title: Periplasmic trehalase from Escherichia coli--characterization and immobilization on spherisorb.
abstract: 1. Trehalase was partially purified from Escherichia coli and characterized. The Km for trehalose was 0.78 mM, the pH optimum 5.5 and the temperature optimum 30 degrees C. 2. Trehalase represented approximately 50% of the total protein released by osmotic shock. The preparation was free of nonspecific carbohydrate hydrolases, which act on sucrose, galactose and maltose, permitting trehalose determination in biological samples, such as insect hemolymph and free cell extracts among others. 3. The enzyme was stable in 50 mM maleate buffer, pH 6.2, at -8 degrees C for at least 6 months and could be used to determine trehalose in the range of 6 to 30 nmol. 4. Immobilization of the enzyme was achieved by covalent linkage to spherisorb-5NH2 (spherical silica gel). Retention of total catalytic activity averaged 32%. 5. The reactor, stored for one month at -5 degrees C, retained 98% of its initial immobilized activity. 6. This immobilized form of the enzyme could be used routinely for specific determinations of trehalose.
authors.affiliation: Departamento de Bioquímica, Universidade Federal do Rio de Janeiro, Brasil.
authors.forename: C F
authors.initials: CF
authors.lastname: Tourinho-dos-Santos
authors.forename: N
authors.initials: N
authors.lastname: Bachinski
authors.forename: V M
authors.initials: VM
authors.lastname: Paschoalin
authors.forename: C L
authors.initials: CL
authors.lastname: Paiva
authors.forename: J T
authors.initials: JT
authors.lastname: Silva
authors.forename: A D
authors.initials: AD
authors.lastname: Panek
mesh_terms: D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004789:Enzyme Activation; D004800:Enzymes, Immobilized / Q000302:isolation & purification* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology*; D006358:Hot Temperature; D058428:Silica Gel; D012822:Silicon Dioxide; D013997:Time Factors; D014198:Trehalase / Q000302:isolation & purification* / Q000378:metabolism; D014199:Trehalose / Q000032:analysis
chemical_list: D004800:Enzymes, Immobilized; D058428:Silica Gel; D012822:Silicon Dioxide; D014199:Trehalose; D014198:Trehalase