Conformation of Myoglobin‐Poly(Ethyl Ethylene Phosphate) Conjugates Probed by SANS: Correlation with Polymer Grafting Density and Interaction

Abstract

One can take advantage of the influence of a polymer conjugated with a protein to control the thermal stability and the deployment of the protein. Here, the structural properties are reported of the protein–polymer conjugate myoglobin (Mb)‐poly(ethyl ethylene phosphate) (PEEP) in the native and unfolded conformations, in order to understand the respective roles of the protein and of the polymer size in the stability of the conjugate. The effect is also investigated of the grafting density of the linear biodegradable polyphosphoesters covalently attached to the protein. It is observed that, while the conjugation process at room temperature does not modify the secondary and tertiary structure of the Mb, the unfolding process, as a function of temperature, depends on the grafting density. Small angle neutron scattering reveals that, at room temperature, conjugation does not alter the size of the native protein and that the thickness of the polymer shell around the protein increases as a function of grafting density and of polymer molecular weight. The denatured form of all conjugates is described by an unfolded chain and a correlation length due to the presence of local stiffness.

How to Characterize the Protein Structure and Polymer Conformation in Protein‐Polymer Conjugates – a Perspective

“…For both proteins, scattering data revealed a picture of a conjugate with a compact protein core surrounded by a polymer shell. [ 106 ] The size and shape of the conjugate could be influenced by the grafting density and molecular weight of attached polymer chains. For BSA, conjugation led to a shape‐shift from a prolate ellipsoid to a more globular shape.…”

“…[ 106 ] It needs to be mentioned that Mb, which is a rather small protein compared to BSA, was less stable in the conjugate form compared to its native form under similar conditions. [ 106 ]…”

“…Russo and coworkers focused on the determination of the polymer arrangement around a protein and the influence of polymer conjugation on protein flexibility and stability at denaturing temperature. [106][107][108] Two different proteins were analyzed, BSA and myoglobin (Mb), a smaller 𝛼-helical protein, to investigate the impact of protein size and secondary structure on the conjugate stability. The polymer of interest was poly(ethyl ethylene phosphate) (PEEP), a biodegradable, temperature-responsive polymer with a transition temperature of ≈60 °C.…”

Poly(ethylene glycol)-Based Surfactant Reduces the Conformational Change of Adsorbed Proteins on Nanoparticles

“…Further studies suggested that the presence of PEG not only reduces the overall unspecific protein adsorption in blood plasma but can also promote the enrichment of particular proteins. , These specific proteins would play a critical role in triggering the highly desired stealth effect. Although functionalization with PEG is currently the most commonly used strategy, new polyphosphoester (PPE) polymers such as poly­(ethyl ethylene phosphate) are emerging as a biodegradable alternative. , A few studies demonstrated that PPE conjugates exhibit similar protein adsorption patterns to those with PEG . Other examples are cylindrical brush polymers such as polysarcosines, which are suitable carriers that extend in vivo circulation times and show high biocompatibility .…”

“…Although functionalization with PEG is currently the most commonly used strategy, new polyphosphoester (PPE) polymers such as poly(ethyl ethylene phosphate) are emerging as a biodegradable alternative. 8 , 9 A few studies demonstrated that PPE conjugates exhibit similar protein adsorption patterns to those with PEG. 10 Other examples are cylindrical brush polymers such as polysarcosines, which are suitable carriers that extend in vivo circulation times and show high biocompatibility.…”