A recoverable and thermoresponsive polymer-protein bioconjugate is synthesized and employed in the purification of protein with free sulfhydryl groups. Initiator with disulphide was modified on the cysteine residue of the target protein. Poly(N-isopropylacrylamide) exhibiting a lower critical solution temperature was grown from the protein. The resulting protein-polymer conjugate was successfully thermoprecipitated and separated from other proteins. The approach was demonstrated with bovine serum albumin with the recycling yield of 76.4%. Enzyme activity test with papain verified the reversible polymer modification protected protein under extreme environments without affecting the functionality of the protein. This study implies the favorable potential of chemo-selective enriching and purification of proteins.

Efficient strategies for enriching and separating proteins are important and challenging for membrane proteomics. Many existing methods are caught in the dilemma of preserving maximal membrane proteins while avoiding the contamination of cytoplasmic proteins and organelles. Here, we report a polymer anchoring strategy for the selective preparation of membrane proteins through cell surface-initiated atom transfer radical polymerization. The cytocompatible polymerization strategy enables thermoresponsive poly(N-isopropylacrylamide) (pNIPPAm) chains to be grown from a specific protein on the surface of living cells. The polymer tagged membrane protein could be easily separated and enriched by thermoprecipitation. This method led to the identification of 1825 proteins of which 1036 (71.7%) were specific membrane proteins in E. coli. The separated proteins were identified by 2-DE and mass spectrometry. Among the 12 protein spots from the gel slice, eight were identified as outer membrane proteins. The described strategy opens up a new avenue for membrane protein enrichment and separation and may expedite the future development of membrane proteomics.

Capsid protein of porcine circovirus type 2 (PCV2) is an ideal subunit vaccine candidate for the postweaning multisystemic wasting syndrome. In this study, mannan-mediated targeting of PCV2ΔCap42‑233 protein to antigen presenting cells (APCs) was investigated for the development of PCV2 vaccine. Mannan was attached to PCV2ΔCap42‑233 protein via an acid sensitive Schiff base reaction. The mannosylated protein was endowed with the capacity to target the mannose receptor on APCs as well as the ability of controlled release of the antigen in the acidic condition of the lysosome. Finally, the immune response of mannosylated PCV2ΔCap42‑233 protein in mice was evaluated. The mannosylated protein exhibited the ability to stimulate humoral immune response and enhance the immunity. Thus, acid-sensitive and APCs-targeting mannosylated PCV2ΔCap42‑233 protein represents a promising candidate for the potential commercial application as an efficient vaccine against porcine circovirus.