Covalent linkage of alkalothermophilic catalase onto functionalized cellulose

Abstract

Catalase from a thermophilic bacterium belonging to the genus Geobacillus was purified and covalently immobilized onto a functionalized polymer via a spacer with an objective to improve its kinetic and biochemical properties. This is the first report on the purification and immobilization of catalase from the genus Geobacillus. The tetrameric catalase of about 221 kDa was successfully purified using a multistep purification strategy. A shift in pH and temperature optima from 8.0 to 9.0 and 55 °C to 60 °C, respectively was recorded after covalent binding of catalase onto the functionalized matrix. The kinetic constants i.e. K_m, V_max, K_cat and K_cat/K_m were found to be 1.2 mM, 4.43 × 10⁶ IU, 6.3 × 10⁵ s⁻¹ and 5.25 × 10⁸ s⁻¹ M⁻¹ for free, and 1.8 mM, 4.01 × 10⁶ IU, 5.9 × 10⁵ s⁻¹ and 3.20 × 10⁸ s⁻¹ M⁻¹ for immobilized catalase, respectively. The ease of binding of alkalothermophilic catalase from a novel isolated bacterium G. extremocatsoochus sp. nov., MTCC 5873 onto a low cost activated cellulose support demonstrated enhanced pH and thermal stability as compared to its free counterpart. This immobilized catalase preparation with improved characteristics has good potential for diverse applications. The present findings provide valuable information on how to tailor enzymes and supported polymer matrices to improve the performance of a biocatalyst.