Protein-protein interactions for solutions of an IgG1 molecule were quantified using static light scattering (SLS) measurements from low to high protein concentrations (c). SLS was used to determine second osmotic virial coefficients (B) at low c, and excess Rayleigh profiles (R/K vs. c) and zero-q structure factors (S) as a function of c at higher c for a series of conditions (pH, sucrose concentration, and total ionic strength [TIS]). Repulsive (attractive) interactions were observed at low TIS (high TIS) for pH 5 and 6.5, with increasing repulsions when 5% w/w sucrose was also present. Previously developed and refined coarse-grained antibody models were used to fit model parameters from B versus TIS data. The resulting parameters from low-c conditions were used as the sole input to multiprotein Monte Carlo simulations to predict high-cR/K and S behavior up to 150 g/L. Experimental results at high-c conditions were quantitatively predicted by the simulations for the coarse-grained models that treated antibody molecules as either 6 or 12 (sub) domains, which preserved the basic shape of a monoclonal antibody. Finally, preferential accumulation of sucrose around the protein surface was identified via high-precision density measurements, which self-consistently explained the simulation and experimental SLS results.
Arginine has been used extensively as an excipient in the formulation development of protein-based biopharmaceuticals. We investigate the role of arginine in suppressing protein aggregation and its mechanism by using bovine serum albumin as a model system. By using sedimentation velocity and other analytical techniques, we show that the use of arginine inhibits temperature-induced aggregation of the protein. We use fluorescence correlation spectroscopy and other spectroscopic techniques to show that arginine inhibits accumulation of partially folded intermediates, potentially involved in the aggregation process. The hydrodynamic radii of the protein in its native, unfolded, and intermediate states have been determined using fluorescence correlation spectroscopy at single-molecule resolution. A possible mechanism of the effects of arginine and its role as an aggregation suppressor has been discussed.
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