Investigation of the operational stabilities and kinetics of glucoamylase immobilized on alkylamine derivatives of titanium(IV)-activated porous inorganic supports |
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Authors: | JMS Cabral JF Kennedy JM Novais |
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Institution: | 2. Laboratório de Engenharia Bioquímica, Instituto Superior Técnico, Universidade Técnica de Lisboa, 1000 Lisbon, Portugal;1. Research Laboratory for the Chemistry of Bioactive Carbohydrates and Proteins, Department of Chemistry, University of Birmingham, P.O. Box 363, Birmingham, B15 2TT, UK |
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Abstract: | Glucoamylase (exo-1,4-α-d-glucosidase, EC 3.2.3.1) was coupled to several porous silica matrices by an improved metal-link/chelation process using alkylamine derivatives of titanium(IV)-activated supports. In order to select the titanium activation procedure which gave stable enzyme preparations, long-term stability tests were performed. The immobilized glucoamylase preparations, in which the carrier was activated to dryness with a 15% w/v TiCl4 solution, displayed very stable behaviour, with half-lives of ~60 days. The optimum operating conditions were determined for these preparations. There are significant differences between the behaviour of the immobilized enzyme and the free enzyme. The apparent Km increased on immobilization due to diffusional resistances. The pH optimum for the immobilized preparation showed a slight shift to acid pH relative to that of the soluble enzyme. Also, the optimum temperature descreased to 60°C after immobilization. In order to test Michaelis-Menten kinetics at high degrees of conversion, time-course analysis of soluble starch hydrolysis was performed. It was observed that simple Michaelis-Menten kinetics are not applicable to the free/immobilized glucoamylase-starch system at high degrees of conversion. |
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Keywords: | Chemical kinetics starch hydrolysis glucoamylase EC 3 2 3 1 immobilization alkylamine derivatives titanium(IV)-activated inorganic supports |
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