Balancing Enzyme Encapsulation Efficiency and Stability in Complex Coacervate Core Micelles

[Image: see text] Encapsulation of charged proteins into complex coacervate core micelles (C3Ms) can be accomplished by mixing them with oppositely charged diblock copolymers. However, these micelles tend to disintegrate at high ionic strength. Previous research showed that the addition of a homopolymer with the same charge sign as the protein improved the stability of protein-containing C3Ms. In this research, we used fluorescence correlation spectroscopy (FCS) and dynamic light scattering (DLS) to study how the addition of the homopolymer affects the encapsulation efficiency and salt stability of the micelles. We studied the encapsulation of laccase spore coat protein A (CotA), a multicopper oxidase, using a strong cationic-neutral diblock copolymer, poly(N-methyl-2-vinyl-pyridinium iodide)-block-poly(ethylene oxide) (PM2VP(128)-b-PEO(477)), and a negatively charged homopolymer, poly(4-styrenesulfonate) (PSS(215)). DLS indeed showed an improved stability of this three-component C3M system against the addition of salt compared to a two-component system. Remarkably, FCS showed that the release of CotA from a three-component C3M system occurred at a lower salt concentration and over a narrower concentration range than the dissociation of C3Ms. In conclusion, although the addition of the homopolymer to the system leads to micelles with a higher salt stability, CotA is excluded from the C3Ms already at lower ionic strengths because the homopolymer acts as a competitor of the enzyme for encapsulation.