Screening and Characterization of Affinity Peptide Tags Specific to Polystyrene Supports for the Orientated Immobilization of Proteins

摘要

Dodecapeptides that exhibit a high affinity specific to a polystyrene surface (PS-tags) were screened using an Escherichia coli random peptide display library system,and the compounds were used as a peptide tag for the site-specific immobilization of proteins.The various PS-tags obtained after 10 rounds of biopanning selection were mainly composed of basic and aliphatic amino acid residues,most of which were arranged in close proximity to one another.Mutant-type glutathione 5-transferases (GSTs) fused with the selected PS-tags,PS19 (RAFIASRRIKRP) and PS23 (AGLRLKKAAIHR) at their C-terminus,GST-PS 19 and GST-PS23,when adsorbed on the PS latex beads had a higher affinity than the wild-type GST,and the specific remaining activity of the immobilized mutant-type GSTs was approximately 10 times higher than that of the wild-type GST.The signal intensity detected for GST-PS 19 and GST-PS23 adsorbed on hydrophilic and hydrophobic PS surfaces using an anti-peptide antibody specific for the N-terminus peptide of GST was much higher than that for the wild-type GST.These findings indicate that the mutant-type GSTs fused with the selected peptide tags,PS19 and PS23,could be site-specifically immobilized on the surface of polystyrene with their N-terniinal regions directed toward the solution.Thus,the selected peptide tags would be useful for protein immobilization in the construction of enzyme-linked immunosorbent assay (ELISA) systems and protein-based biochips.