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To develop the technology for CO2 sequestration, the free and immobilized carbonic anhydrase treatment was performed. The amount of dissolved CO2 was increased by 3-fold after enzyme treatment of 5 ppm, and the amount of calcium carbonate increased linearly as the CaCl2 was increased to 100 mM. To immobilize the carbonic anhydrase, an aluminum oxide carrier surface was functionalized with octadecyltrichlorosilane, 1H,1H,2H,2H-perfluoroctyltriethoxysilane, 3-aminopropyltriethoxysilane, 3-mercaptopropyl triethoxysilane, chlorotrimethylsilane 3-cyanopropyltrichlorosilane or 3-glycidyloxypropyltriethoxysilane. The best effect among these substances was revealed with 3-aminopropyltriethoxysilane; the efficacy of the immobilized carbonic anhydrase was at over 99%, and even when it was repeatedly used up to 40 times, the efficacy of the immobilized enzyme was more than 80%. The optimal conditions for the immobilized enzyme were 60oC and pH 7.0.

The gene encoding 1,4-β-endoglucanase KRICT PC-001 (1.722 bp; 573 aa) was revealed by a full-length sequence analysis of the bacterium Paenibacillus terrae HPL-003 (KCTC 11987BP) isolated from the soil on Gara Mountain in Korea (CP003107). The molecular weight of the enzyme was estimated as 62 kDa by SDS-PAGE analysis. Endoglucanase KRICT PC-001 contains a catalytic domain of the glycosyl hydrolase family 5 (GH5) and a carbohydrate-binding module X2 (CBM X2). The soluble fraction of the recombinant protein was overexpressed with the pGEX-fusion vector in Escherichia coli BL21 (DE3) and purified using glutathione S-transferase column. The recombinant protein was digested with Factor Xa and eluted from a benzamidine column. Carboxymethyl cellulose was used as a substrate for the enzyme activity analysis. Endoglucanase KRICT PC-001 exhibited a maximum specific activity of 589 U/mg of protein at a temperature of 50°C and pH 5.0.