Final Technical/Scientific Report: Commodity Scale Thermostable Enzymatic Transformations

Lalonde, James J. ; Davison, Brian

Altus Biologics (United States)

关键词: Isomerase;    Agglomeration;    Starch;    Corn;    Fructose;   

DOI  :  10.2172/813693
RP-ID  :  DOE/CH11006
RP-ID  :  FC07-99CH11006
RP-ID  :  813693

美国|英语

来源: UNT Digital Library

【 摘 要 】

The conversion of corn starch to high fructose corn-syrup sweetener is a commodity process, producing over 3 billion kg/y. In the last step of the process, an enzyme catalyst is used to convert glucose to the much sweeter sugar fructose. Due to incomplete conversion in the last step, the syrup must be purified using a chromatographic separation technique, which results in equal quantities of water being added to the syrup, and finally the water must be evaporated (up to 1 lb of water/lb of syrup). We have estimated the energy requirement in the evaporation step to be on the order of 13 billion BTU's/y. This process inefficiency could be eliminated if a thermostable form of glucose isomerase (GI), the enzyme catalyst used in the final step, was developed. Our chosen strategy was to develop an immobilized form of the enzyme in which the protein is first crystallized and then chemically cross-linked to form an insoluble particle. This so-called cross-linked enzyme crystal (CLE C(reg. sign)) technology had been shown to be a powerful method for enzyme stabilization for several other protein catalysts. In this work we have developed more than 30 CLEC preparations of glucose isomerase and tested them for activity and stability. We found these preparations to be highly active, with a 10-50 fold rate per gram of catalyst increase over existing commercial catalysts. The initial rates were also higher at higher temperatures as expected, however the efficiency of the CLEC GI preparations unexpectedly rapidly decreased to a low constant value with use at the higher temperatures. At this point, the source of this activity loss is unclear, however during this loss, the catalyst is found to form a solid mass indicating either breakage of the chemical cross-links or simple aggregation of the particles. It is likely that the increased mass transfer resistance due to this agglomeration is a major component of the activity loss. This research suggests that one potentially beneficial outcome could be the reconfiguration of catalyst columns using these highly active catalyst preparations with inerts to prevent agglomeration. As a result of this work, methods for the preparation of highly active immobilized glucose isomerase preparations were developed along with test methods that are predictive for the stability of these preparations. This research has been conducted as a team effort. The enzyme is produced using Genencor's glucose isomerase protein and the stabilized form has been prepared using Altus' CLEC technology. ORNL has provided bioprocess engineering and testing expertise, and Cargill, Inc. and Genencor have supplied critical technical consultation and economic assessment.