Influence of the immobilisation process on the activity of β-galactosidase bound to nylon membranes grafted with glycidyl methacrylate Part 2.: Non-isothermal behaviour

beta -Galactosidase was immobilised via diazotisation or condensation on nylon membranes, chemically grafted with glycidyl methacrylate (GMA) and having identical physical transport properties, such as hydraulic and thermoosmotic permeabilities. Immobilisation by diazotisation involves the binding of the enzyme to the activated membranes through the tyrosine residues of the macromolecule, while immobilisation by condensation occurs through arginine residues. The performance of both membranes types was studied as a function of substrate concentration, average temperature and temperature gradients in a bioreactor operating under non-isothermal conditions. Both membrane types exhibit apparent K-m values of the immobilised form higher than that of the free enzyme. Under non-isothermal conditions the apparent K-m of the immobilised beta -galactosidase approached values more close to that of the, free enzymes recovering in this way the loss of affinity consequent to the immobilisation process. Moreover, membranes obtained via diazotisation resulted more efficient than those obtained via condensation, showing higher percentage activity increases with respect to all the experimental variables studied. This behaviour has been attributed to the different immobilisation technique, since the multipoint enzyme attachment through the arginine residues of the macromolecule affects the catalytic structure of the catalytic site more than the attachment by diazotisation. Activity increases under non-isothermal conditions were found to decrease with the increases of substrate concentration and average temperature. The values of the yield increases with 1 degreesC of temperature difference across both catalytic membranes confirm the useful employment of the technology of the non-isothermal bioreactors in productive biotechnological processes. (C) 2001 Published by Elsevier Science B.V.