RESUMO
Thermotoga maritima exo-b-fructosidase (BfrA) secreted by a recombinant Pichia pastoris strain was optimall y immobilised on GlyoxylSepharose CL 4B using the Rational Design of Immobilised Derivatives(RDID) strategy. Covalent attachment of the N-glycosylated BfrA on to the activated support at pH 10allowed total recovery of the loaded enzyme and its activity. The immobilisation process caused no variationin the catalytic properties of the enzyme and allowed further enhancement of the thermal stability.Complete inversion of cane sugar (2.04 M) in a batch stirred tank reactor at 60 C was achieved with aproductivity of 22.2 g of substrate hydrolysed/gram of biocatalyst/hour. Half-life of the immobilisedenzyme of 5 days at 60 C was determined in a continuously operated fixed-bed column reactor. Ourresults promote the applicability of the BfrA-immobilised biocatalyst for the complete hydrolysis of concentratedsucrose solutions under industrial conditions, especially at a high reaction temperature(AU)
