Protein–polymer
bioconjugate self-assembly has attracted
a great deal of attention as a method to fabricate protein nanomaterials
in solution and the solid state. To identify protein properties that
affect phase behavior in protein–polymer block copolymers,
a library of 15 unique protein-b-poly(N-isopropylacrylamide) (PNIPAM) copolymers comprising 11 different
proteins was compiled and analyzed. Many attributes of phase behavior
are found to be similar among all studied bioconjugates regardless
of protein properties, such as formation of micellar phases at high
temperature and low concentration, lamellar ordering with increasing
temperature, and disordering at high concentration, but several key
protein-dependent trends are also observed. In particular, hexagonal
phases are only observed for proteins within the molar mass range
20–36 kDa, where ordering quality is also significantly enhanced.
While ordering is generally found to improve with increasing molecular
weight outside of this range, most large bioconjugates exhibited weaker
than predicted assembly, which is attributed to chain entanglement
with increasing polymer molecular weight. Additionally, order–disorder
transition boundaries are found to be largely uncorrelated to protein
size and quality of ordering. However, the primary finding is that
bioconjugate ordering can be accurately predicted using only protein
molecular weight and percentage of residues contained within β
sheets. This model provides a basis for designing protein–PNIPAM
bioconjugates that exhibit well-defined self-assembly and a modeling
framework that can generalize to other bioconjugate chemistries.
Nanoparticle (NP) carriers provide new opportunities for controlled delivery of drugs, and have potential to address challenges such as effective oral delivery of insulin. However, due to the difficulty of efficiently loading insulin and other proteins inside polymeric NPs, their use has been mostly restricted to the encapsulation of small molecules. To better understand the processes involved in encapsulation of proteins in NPs, we study how buffer conditions, ionic chelation, and preparation methods influence insulin loading in poly (lactic-co-glycolic acid)–b–poly(ethylene glycol) (PLGA-PEG) NPs. We report that, although insulin is weakly bound and easily released from the NPs in the presence of buffer ions, insulin loading can be increased by over 10-fold with the use of chelating zinc ions and by the optimization of the pH during nanoprecipitation. We further provide ways of changing synthesis parameters to control NP size while maintaining high insulin loading. These results provide a simple method to enhance insulin loading of PLGA-PEG NPs, and provide insights that may extend to other protein drug delivery systems that are subject to limited loading.
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