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(626b) Determination of the Conformation of Protein-Conjugated Poly(ethylene glycol) Chains by Small-Angle Neutron Scattering

Pai, S. S., Carnegie Mellon University
Przybycien, T. M., Carnegie Mellon University
Tilton, R. D., Carnegie Mellon University

Covalent conjugation of poly(ethylene glycol) (PEG) to proteins has proven to be an effective strategy to increase in vivo circulation times of protein therapeutics, with consequent improvements in pharmaceutical efficacy. Recent work indicates that PEG conjugation may also significantly improve the preservation of protein native structure and bioactivity during sustained release from biodegradable drug delivery depots. Understanding the configuration of PEG-conjugated proteins is necessary to interpret the effect of PEG conjugation on renal clearance rates and steric protection of circulating proteins in vivo, as well as effects on PEG-protein adsorption and associated denaturation in sustained-release delivery devices. Dynamic light scattering studies have suggested that PEG-protein conjugates adopt a dumbbell configuration, with a relatively unperturbed PEG random coil adjacent to the globular protein. This is inconsistent with the conventionally presumed shroud model where PEG chains are proposed to ?wrap? around the protein, creating a shielding effect. Hypothesizing that PEG configurational entropy disfavors the shroud model, we employed Small-Angle Neutron Scattering (SANS) to distinguish between the dumbbell model and the shroud model for chicken egg lysozyme covalently attached to a single 20 kDa PEG chain. The SANS contrast variation technique was used to isolate the PEG portion of the conjugate. SANS data are well described by the dumbbell model, but are inconsistent with the shroud model. Investigations of the effect of the size of the PEG chain relative to the protein are currently underway to determine if the shroud model becomes more likely with increasing PEG size.