A NOVEL CLASS OF PROTEIN-POLYMER CONJUGATES WITH BIODEGRADABLE POLYPHOSPHOESTERS: SYNTHESIS, BIOCHEMICAL, AND BIOPHYSICAL EVALUATION

Polyphosphoesters (PPEs) are a novel class of polymers, which present high potential in the substitution of the gold standard poly(ethylene glycol) (PEG) in the biomedical field. We present the synthesis, biochemical and biophysical evaluation of a novel set of protein-polymer conjugates made by the protein myoglobin and PPEs with various hydrophilicity and molar mass. The polymers were synthesised via a ring-opening polymerisation procedure. Their hemocompatibility in human blood and biodegradability in human fluids was assessed. The polymers were further used for the bioconjugation reaction with the protein myoglobin. The samples were purified, characterised, and their biochemical properties in solution were evaluated. In particular, we studied the conformation, the residual activity and the thermal stability of the protein, observing a ladder of properties linked to the polymer hydrophilicity. Moreover, the more hydrophilic polymers showed a protective action toward thermal-induced aggregation and proteolytic enzymes’ attack. In the second part of the thesis, the biophysical properties of the conjugates in the dry form were tested by the complementary approach of neutron scattering spectroscopy and Differential Scanning Calorimetry. We observed the samples’ dynamics at different time scales, with the aim to correlate it to the structure and the functionality of the system. The studies performed permitted us to draw a microscopic picture of our samples in the dry form, shedding light on the establishment of interactions among the protein and the polymer chains. The results will help the design of future conjugates.