Structural Response of Bovine Growth Hormone to Dead-Ended Ultrafiltration

Abstract

Bovine growth hormone (bGH) was used as a model pharmaceutical protein to assess protein structural stability during dynamic ultrafiltration. Two popular membrane materials of different wettabilities, poly(ether sulfone) (PES) and regenerated cellulose (RC), are exposed to acidic and basic bGH solutions in a dead-ended filtration. After a three-fold concentration run, samples of the feed, permeate, and retentate are subjected to a series of spectroscopic and physical analytical techniques, permitting quantitative characterization of the secondary and tertiary structure of bGH as well as the aggregation state of the bGH in these streams. At pH 4, where the monomeric form predominates in the feed, bGH loses helix and gains sheet content as it passes through the membranes. At pH 9, where the feed contains mainly dimers, bGH in the permeate is mainly monomeric and also gains sheet content at the apparent expense of helix content. Tertiary structure analyses suggest that passage through the membranes induces bGH to adopt a more compact form than the bGH in either the feed or retentate streams. At both pHs 4 and 9, the less wettable PES membrane induces a greater structural change in the permeating bGH than does the more wettable RC membrane. Alkaline pH also tends to preserve the structure of bGH in the retentate streams, perhaps reflecting the extensive dimerization. Our results suggest that the choice of filtration conditions is critical in preserving protein structure and, hence, bioactivity.