DNA-directed trypsin immobilization on a polyamidoamine dendrimer-modified capillary to form a renewable immobilized enzyme microreactor.
A novel type of trypsin capillary microreactor was developed based on a DNA-directed immobilization (DDI) technique applied to a fused-silica capillary modified with polyamidoamine (PAMAM) dendrimers. Trypsin binding to the inner wall of the capillary was confirmed by confocal laser scanning microscopy. The properties of the trypsin-DNA conjugated, PAMAM-modified capillary microreactor were investigated by monitoring hydrolysis of Nα-benzoyl-L-arginine ethyl ester. Through the hybridization and dehybridization of the DNA, the inner wall of the capillary functionalized with trypsin can be regenerated, thus indicating the renewability of this enzyme microreactor. In addition, these results demonstrated that introduction of PAMAM enabled higher amounts of trypsin to be immobilized, markedly improving the enzymolysis efficiency, compared with traditional modified capillaries. The digestion performance of the trypsin capillary microreactor was further evaluated by digesting cytochrome C, and a peptide numbers of 8, and a sequence coverage of 59% were obtained. This renewable and efficient immobilized trypsin capillary microreactor combines advantages of both DDI technology and PAMAM, and is potentially adaptable to high-throughput enzyme assays in biochemical and clinical research.
['Wu|Nan|N|', 'Wang|Siming|S|', 'Yang|Ye|Y|', 'Song|Jiayi|J|', 'Su|Ping|P|', 'Yang|Yi|Y|']
D000818:Animals D002417:Cattle D004247:DNA D050091:Dendrimers D004800:Enzymes, Immobilized D006868:Hydrolysis D007700:Kinetics D008958:Models, Molecular D009690:Nucleic Acid Conformation D011073:Polyamines D014357:Trypsin
Trypsin DNA-directed immobilization Capillary enzyme microreactor