Formation of molecularly imprinted polymers with trypsin activity

• Main Authors: Wen-Ru Chen, 陳玟茹
• Other Authors: Dar-Fu Tai
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/4gparc

碩士 === 國立東華大學 === 化學系 === 95 === Enzymes exhibit catalytic activity and high selectivity. They are widely applicable in various field. However, industrial applications of enzymes are still limited due to their instability and cost. Additionally, it is difficult to obtain enzyme at high purity. Molecularly imprinted polymers (MIPs) are usually specialized with recognition ability or catalytic activity. We attempted to create the polymers with trypsin activity using molecular imprinting. The first goal is to produce polymeric materials with stereo-recognition site toward trypsin on gold surface. In the presence of trypsin as template, the resulting porous MIPs contained complementary functional group and the size, shape memory of trypsin. Proficiently, the trypsin absorpted MIPs was able to catalyze the hydrolysis of N-Benzoyl-L-Arg-ethylester (BAEE). Second, based on the serine protease’s mechanism, the active sites of trypsin are imitated to construct the artificial enzyme. Three amino acids were modified. Their corresponding acrylates were served as major functional monomers. However, substrate analogues were used as template to generate the cavities on gold or fiber matrix by heat induced polymerization. The created MIPs-cavity demonstrated similar activity as trypsin and hydrolyzed the target substrate BAEE. The kinetic analysis show the parameters of MIPs had Vmax = 5x10-8 [Mmin-1], kcat = 8x10-5 [min-1], Km = 5x10-5 [M], kcat / Km = 1.6 [min-1M-1].