Preparation of biodegradable and thermoresponsive enzyme-polymer conjugates with controllable bioactivity via RAFT polymerization

The preparation of biodegradable and thermoresponsive enzyme-polymer bioconjugates with controllable enzymatic activity via reversible addition-fragmentation chain transfer (RAFT) polymerization and amidation conjugation reaction is presented. A new 2-mercaptothiazoline ester functionalized RAFT agent with intra-disulfide linkage was synthesized and used as chain transfer agent (CTA) to generate a biocompatible homopolymer, poly(ethyleneglycol) acrylate (polyPEG-A) and a thermoresponsive copolymer of poly(ethyleneglycol) acrylate with di(ethyleneglycol)ethyl ether acrylate [poly(PEG-A-co-DEG-A)]. These biodegradable and thermoresponsive polymers were then conjugated to the surface of glucose oxidase (GOx) under mild condition to afford the biodegradable and thermoresponsive enzyme-polymer conjugates. Cleavage of the polymer chains from the GOx surface obviously recovered the enzymatic activity. The thermoresponsive test of GOx-poly(PEG-A-co-DEG-A) revealed that the bioconjugate exhibited regular enzymatic activity fluctuation upon the temperature change below or above the lower critical solution temperature (LCST). The as-prepared enzyme-polymer conjugates were also characterized using 1H NMR, UV-vis spectroscopy, polyacrylamide gel electrophoresis (PAGE) and biocatalytic activity tests. These smart enzyme-polymer conjugates would envision promising applications in biotechnology and biomedicine.