Immobilization of AaCel9a endoglucanase on chitosan macrobeads

Cellulase enzyme has shown their potential application in different industry. cellulase immobilization is one of the different methods for enzymatic stabilization. An advantage of immobilization is enzymatic reusability, which have an economical advantage for enzyme using in industry. Properties of Chitosan as a support for enzyme immobilization are always considerable. Due to its unique biological properties such as biocompability, biodegradability and non-toxicity, chitosan is an attractive support for immobilization. In this investigation Aa-cel9A endoglucanase gene was cloned in pET28 () expression vector. Sequencing result had been proved gene cloning in vector. Then the constructed vector was transformed to Eshershia.Coli (BL21) cells and enzyme production was induced. The result obtained from SDS-PAGE analysis and enzymatic assay showed the recombinant protein has been expressed and protein purification was done with Ni-NTA column. Chitosan macrobeads were prepared by precipitation procedure. After immobilization of enzyme with glutaraldehyde as linker, enzyme immobilization has been proved with FTIR and Bradford analysis. The obtained result showed optimum condition for covalent immobization on support are 0.7% of glutaraldehyde concentration and sodium phosphate buffer with pH 7. Bradford analysis and enzymatic activity assay have proved 85% of enzyme molecules immobilized on support.