Effect of Polymer Chain Density on Protein–Polymer Conjugate Conformation

Abstract

Many biomedical applications employ covalent attachment to synthetic polymers to enhance the efficiency of proteins or other therapeutically active molecules. We report here the impact of polymer conjugation on the structural and thermal stability of a protein model, the bovine serum albumin, using a variable number of linear biodegradable polyphosphoesters, which were covalently tethered to the protein. We observed that BSA’s secondary structure measured by circular dichroism is independent of the conjugation. Small-angle neutron scattering, however, reveals a change from ellipsoid to globular shape of the whole complex arising from a slight compaction of the protein core and an increase of the polymer’s radius of gyration as a function of the grafting polymer density. In particular, we highlight a gradual change of the polymer conformation around the protein and elongation of the semimajor dimension of the ellipsoidal protein. Our results will contribute to the description of biophysical characteristics of a new class of biologically relevant protein–polymer conjugates.

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

“…The spectra show how the helicoidal structure is conserved in all samples and that, at room temperature, the conjugation does not alter the native Mb folding (Figure S4e, Supporting Information), as previously found for the BSA protein with the same polymer conjugation. [ 9 ]…”

“…Since the fluorescence signal is sensitive to the microenvironments, it is plausible that the conjugation increases the exposure of tryptophan to the solvent rather than producing a screening effect as observed for the BSA. [ 9 ] In fact, both Trp are present in the α‐helix of the Mb near the N‐terminus of the protein and more likely to be exposed to the solvent. In particular, we observe for Mb‐10p and Mb‐20p that an initial decreasing of the λ max (blue shift) is followed by a sudden increase at 50 °C.…”

“…of a particular polymer to its ultimate effect on the biological molecule. [ 5 ] As we stressed in our previous publications, [ 6–9 ] the understanding of the biophysical characteristics, interactions, structure, and dynamical properties of protein–polymer conjugates is fundamental to depict the polymer impact in the protein folding and activity. The goal is to answer the following questions: How does polymer conjugation influence protein folding/unfolding and stability?…”

“…We evidenced a gradual change of the polymer conformation on the protein surface as a function of grafting density, from a compact to open Gaussian chain toward a star‐like shape. [ 9 ] In order to identify general criteria, in the presented work we focused our attention on the structural properties of Myoglobin conjugates. We studied the impact on the size and α‐helix secondary structure of the protein.…”

Insight into Protein–Polymer Conjugate Relaxation Dynamics: The Importance of Polymer Grafting

“…A gradual change of the polymer conformation as a function of grafting density, from a compact to open Gaussian chain toward a star‐like shape was also evidenced. [ 18 ]…”