Stabilization of immobilized l‐arabinose isomerase for the production of d‐tagatose from d‐galactose

Abstract The aim of this work was to develop a stable immobilized enzyme biocatalyst for the isomerization of d ‐galactose to d ‐tagatose at high temperature. l ‐Arabinose isomerase from the hyperthermophilic bacterium Thermotoga maritima (TMAI) was produced as a (His) 6 ‐tagged protein, immobilized on a copper–chelate epoxy support and subjected to several postimmobilization treatments aimed at increasing its operational and structural stability. Treatment with glutaraldehyde and ethylenediamine resulted in a more than twofold increase in the operational stability and in all enzyme subunits linked, directly or indirectly, to the support via covalent bonds. A postimmobilization treatment of the immobilized derivatives with mercaptoethanol for the removal of any remaining copper ions, determined a further increase of the operational biocatalytic activity. Immobilized derivatives subjected to both treatments were used for the bioconversion of 18 g/L d ‐galactose to d ‐tagatose at 80°C in a packed bed reactor in three repeated cycles and showed a better operational stability compared with the literature data. This study shows that a postimmobilization stabilization treatment with glutaraldehyde and ethylenediamine can stabilize the multi‐subunit structure of an enzyme immobilized on a metal‐chelate epoxy support with an increase of its operational stability, results that are not easily achievable with the sole immobilization on epoxy or metal chelate‐epoxy supports in the case of complex multimeric enzymes with geometric incongruence with the support.