Biophysical Society Thematic Meeting - October 13-15, 2015

Biophysics of Proteins at Surfaces: Assembly, Activation, Signaling

Poster Abstracts

12-POS Board 12 Fast Insulin Amyloid Aggregation at the Air-Liquid-Solid Triple Interface Thibaut Frachon 2,1 , Marianne Weidenhaupt 1 , Quentin Le masne 2 , Franz Bruckert 1 . 1 LMGP, Grenoble, France, 2 Eveon, Grenoble, France. With the growing use of therapeutical proteins and the development of medical delivery devices, protein stability becomes a major concern in pharmaceutical industrial processes. Proteins can lose their native conformation by unfolding and reach a new free energy minimum by forming insoluble aggregates. Insulin is one example of such an aggregation-prone protein, which easily forms amyloid fibrils on hydrophobic surfaces under agitation. Within a device containing a protein solution, hydrophobic interfaces are found at the border between hydrophobic materials and liquid and between air and liquid. These different interfaces come into close proximity in protocols, frequently used in industry, involving intermittent wetting. We designed a model experiment, where we observe by optical microscopy the formation of insulin amyloid fibers as a protein solution moves repetitively back and forth in a microfluidic channel. Thioflavin T fluorescence was used to monitor the formation of amyloid deposits and reflection interference contrast microscopy was used to monitor the morphology of the liquid film after receding. The growth of protein aggregates on the surface was characterized. We demonstrate that insulin fibers mainly form in regions of intermittent wetting, but not in regions exposed to hydrodynamic shear stress alone, remaining always wet. This clarifies the role of interfaces in insulin aggregation, showing that wall shear stress alone is insufficient to rapidly trigger amyloid fiber formation. We study the influence of temperature, channel size, fluid velocity, duration of the recession phases and nature of the materials surfaces. Our experiments illustrate how the formation of a triple solid/liquid/air interface in industrial processes may lead to protein instability. The careful design of fluid movement in devices handling protein solutions can therefore improve their stability.

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