title: Improvement of the homogeneity of protein-imprinted polymer films by orientated immobilization of the template.
abstract: A method for preparing homogeneous protein-imprinted polymer films with orientated immobilization of template is described. The template methyl parathion hydrolase (MPH) was modified with a peptide linker (Gly-Ser)(5)-Cys and was immobilized on a cover glass with a fixed orientation via the linker. The activity of the fusion enzyme (MPH-L) was evaluated by determining the product's absorbance at 405 nm (A(405)). Both the free and the immobilized MPH-L showed higher retention of the bioactivity than the wide type enzyme (MPH-W) as revealed by the A(405) values for MPH-L(free)/MPH-W(free) (1.159/1.111) and for MPH-L(immobilized)/MPH-W(immobilized) (0.348/0.118). The immobilized MPH-L also formed a more homogeneous template stamp compared to the immobilized MPH-W. The molecularly imprinted polymer films prepared with the immobilized MPH-L exhibited high homogeneity with low Std. Deviations of 80 and 200 from the CL intensity mean volumes which were observed for batch-prepared films and an individual film, respectively. MPH-L-imprinted polymer film also had a larger template binding capacity indicated by higher CL intensity mean volume of 3900 INT over 2500 INT for MPH-W-imprinted films. The imprinted film prepared with the orientated immobilization of template showed an imprinting factor of 1.7, while the controls did not show an imprinting effect.
authors.affiliation: Key Laboratory of Fermentation Engineering, Ministry of Education, College of Bioengineering, Hubei University of Technology, Wuhan, PR China.
authors.forename: Lijian
authors.initials: L
authors.lastname: Liu
authors.forename: Jingjing
authors.initials: J
authors.lastname: Zheng
authors.forename: Guijie
authors.initials: G
authors.lastname: Fang
authors.forename: Weihong
authors.initials: W
authors.lastname: Xie
mesh_terms: D000595:Amino Acid Sequence; D000906:Antibodies / Q000276:immunology; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005898:Glass / Q000737:chemistry; D007118:Immunoassay; D054802:Molecular Imprinting; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011108:Polymers / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D000906:Antibodies; D004800:Enzymes, Immobilized; D010455:Peptides; D011108:Polymers; D011993:Recombinant Fusion Proteins; D010744:Phosphoric Monoester Hydrolases