Do Protein Molecules Unfold in a Simple Shear Flow?

Protein molecules typically unfold (denature) when subjected to extremes of heat, cold, pH, solvent composition, or mechanical stress. One might expect that shearing forces induced by a nonuniform fluid flow would also destabilize proteins, as when a protein solution flows rapidly through a narrow channel. However, although the protein literature contains many references to shear denaturation, we find little quantitative evidence for the phenomenon. We have investigated whether a high shear can destabilize a small globular protein to any measurable extent. We study a protein (horse cytochrome c, 104 amino acids) whose fluorescence increases sharply upon unfolding. By forcing the sample through a silica capillary (inner diameter 150–180 μm) at speeds approaching 10 m/s, we subject the protein to shear rates dvz/dr as large as ∼2 × 105 s−1 while illuminating it with an ultraviolet laser. We can readily detect fluorescence changes of <1%, corresponding to shifts of <∼0.01 kJ/mol in the stability of the folded state. We find no evidence that even our highest shear rates significantly destabilize the folded protein. A simple model suggests that extraordinary shear rates, ∼107 s−1, would be required to denature typical small, globular proteins in water.