Study On The Immobilization Of Glucose Isomerase And Its Catalytic Properities

Glucose isomerase,also known as xylose isomerase,which is one of key enzymes in the production of high fructose corn syrup process in the food industry had been widely used in the food,beverage,pharmaceutical and other industries.High fructose corn syrup is an ideal source of sugar for modern people to pursue a healthy life.High fructose corn syrup may be divided into three types according to the different content of fructose?42%,55%,90%?:type 42,type 55and type 90.Type 55 is commonly used in food and beverages,but it hadn't be directly produced from glucose.It was mainly concentrated from type 42,with complicated separation process and expensive production cost.As literatures reported,55 type of high fructose corn syrup can be directly prepared with glucose isomerase at temperature above 85?.At the same time,it is important to find a suitable immobilization for the enzyme.In this paper,the immobilization of glucoseisomerasefromrecombinantstrainofE.coli BL21?DE3?/pET28-b-TEGI was studied.In the present study,the immobilization of glucose isomerase using glutaraldehyde cross-linking cell and its granulation preparation was studied.First,effect of cells state of?whole cells and partion broken cells?on the immobilization efficiency were studied.It showed that intact cells could be effectively immobilized,especially in the addition of flocculant.More activity would decrease immobilized with the broken cells,but the operational stability would be improved.The effects of freezing on the immobilized enzyme activity,stability and particle shape was also investigated.The results showed that the immobilized enzyme by frozen treatment took an increased activity.The effects of combined use of flocculants?polyethyleneimine and chitosan?and glutaraldehyde was studied.It showed that enzyme immobilized with the addition of polyethyleneimine was better.The immobilization conditions were further optimized.It showed that the optimal immobilization conditions were as follows with diatomaceous earth addition of 0.6%?w/v?,2%?v/v?polyethyleneiminehe and 0.5%?v/v?glutaraldehyde,at pH 6.0 and room temperature.At the optimal buffer of Na₂HPO₄-NaH₂PO₄ and the optimal immobilization time of1.5-2 h,the activity recovery was reached 80-90%.Finally,the catalytic properties of the enzyme immobilized with polyethyleneiminehe and glutaraldehyde were studied.The results showed that the optimal temperature and pH for the immobilized and free enzyme was 80?and 7.0,respectively.The activity was activated with the addition of Mg²⁺,Fe²⁺,Zn²⁺,Mn²⁺or Co²⁺,and Ba²⁺,Ni²⁺,Ca²⁺or Cu²⁺could inhibit the enzyme activity and stability.The highest conversion was 53.96%.After reaction 10 batches under the optimal conditions,90%of the initial activity was still reserved,and the conversion was all above 46.70%.The immobilized enzyme activity remained unchanged after stored at room temperature for 23 d.