Development of self-cooperative nanochaperones with enhanced activity to facilitate protein refolding.

Regulating protein folding including assisting de novo folding, preventing misfolding and aggregation, and facilitating refolding of proteins are of significant importance for retaining protein's biological activities. Here, we report a mixed shell polymeric micelle (MSPM)-based self-cooperative nanochaperone (self- CO -nChap) with enhanced activity to facilitate protein refolding. This self- CO -nChap was fabricated by introducing Hsp40-mimetic artificial carriers into the traditional nanochaperone to cooperate with the Hsp70-mimetic confined hydrophobic microdomains. The artificial carrier facilitates transfer and immobilization of client proteins into confined hydrophobic microdomains, by which significantly improving self- CO -nChap's capability to inhibit unfolding and aggregation of client proteins, and finally facilitating refolding. Compared to traditional nanochaperones, the self- CO -nChap significantly enhances the thermal stability of horseradish peroxidase (HRP) epicyclically under harsher conditions. Moreover, the self- CO -nChap efficiently protects misfolding-prone proteins, such as immunoglobulin G (IgG) antibody from thermal denaturation, which is hardly achieved using traditional nanochaperones. In addition, a kinetic partitioning mechanism was devised to explain how self- CO -nChap facilitates refolding by regulating the cooperative effect of kinetics between the nanochaperone and client proteins. This work provides a novel strategy for the design of protein folding regulatory materials, including nanochaperones.