The immobilization of proteins on biodegradable polymer fibers via click chemistry.
A facile and efficient method to immobilize bioactive proteins onto polymeric substrate was established. Testis-specific protease 50 (TSP50) was immobilized on ultrafine biodegradable polymer fibers, i.e., (1) to prepare a propargyl-containing polymer P(LA90-co-MPC10) by introducing propargyl group into a cyclic carbonate monomer (5-methyl-5-propargyloxycarbonyl-1,3-dioxan-2-one, MPC) and copolymerizing it with l-lactide; (2) to electrospin the functionalized polymer into ultrafine fibers; (3) to azidize the TSP50, and (4) to perform the click reaction between the propargyl groups on the fibers and the azido groups on the protein. The TSP50-immobilized fibers can resist non-specific protein adsorptions but preserve specific recognition and combination with anti-TSP50. ELISA tests were carried out by using HRP-goat-anti-mouse-IgG(H+L) as secondary antibody and o-phenylenediamine (OPDA)/H(2)O(2) as substrate to detect the combination of immobilized TSP50 with anti-TSP50. The results showed that anti-TSP50 can be selectively adsorbed from its solution onto the TSP50-immobilized fibers in the presence of BSA of as high as 10(4) times concentration. TSP50 immobilized on the fiber and anti-TSP50 combined to the fiber were also quantitatively determined. Anti-TSP50 can be then eluted off from the fiber when pH changes. The eluted fiber can re-combine anti-TSP50 at an efficiency of 75% compared to the original TSP50-immobilized fiber. Therefore, the TSP50-immobilized fibers can be used in the detection, separation, and purification of anti-TSP50. The "click" method can lead to a universal strategy to protein immobilization.
State Key Laboratory of Polymer Physics and Chemistry, Changchun Institute of Applied Chemistry, Chinese Academay of Sciences, Changchun, Jilin 130022, PR China.
Quan
Q
Shi
Xuesi
X
Chen
Tiancheng
T
Lu
Xiabin
X
Jing
D000906:Antibodies / Q000276:immunology / Q000302:isolation & purification; D001672:Biocompatible Materials / Q000138:chemical synthesis / Q000737:chemistry*; D003300:Copper / Q000737:chemistry; D004560:Electricity; D004797:Enzyme-Linked Immunosorbent Assay; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000276:immunology; D006880:Hydroxy Acids; D019344:Lactic Acid / Q000737:chemistry; D008855:Microscopy, Electron, Scanning; D015394:Molecular Structure; D010293:Pargyline / Q000031:analogs & derivatives / Q000737:chemistry; D011108:Polymers / Q000138:chemical synthesis / Q000737:chemistry*; D011422:Propionates / Q000737:chemistry; D011506:Proteins / Q000737:chemistry* / Q000276:immunology; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000276:immunology; D012710:Serum Albumin, Bovine / Q000737:chemistry; D019810:Sodium Azide / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared
D000906:Antibodies; D001672:Biocompatible Materials; D004800:Enzymes, Immobilized; D006880:Hydroxy Acids; D011108:Polymers; D011422:Propionates; D011506:Proteins; C042514:cuprous ion; D012710:Serum Albumin, Bovine; D019344:Lactic Acid; C110034:2,2-bis(hydroxymethyl)-propionic acid; D003300:Copper; D019810:Sodium Azide; D010293:Pargyline; D012697:Serine Endopeptidases; C446379:testis-specific protease 50; C458239:propargyl bromide