Attachment of Proteins to a Hydroxyl-Terminated Surface Eliminates the Stabilizing Effects of Polyols.

Abstract

The physics of proteins interacting with surfaces can differ significantly from those seen when the same proteins are free in bulk solution. As an example, we describe here the extent to which site-specific attachment to a chemically well-defined macroscopic surface alters the ability of several stabilizing and destabilizing cosolutes to modulate protein folding thermodynamics. We determined this via guanidinium denaturations performed in the presence of varying concentrations of cosolutes when proteins were either site-specifically attached to self-assembled monolayers on gold or free in bulk solution. Doing this we found that the extent to which guanidinium (a destabilizing Hofmeister cation), sulfate (a stabilizing Hofmeister anion), and urea (a neutral denaturant) alter the folding free energy remains indistinguishable whether proteins are surface-attached or free in bulk solution. In sharp contrast, however, neutral osmolytes sucrose and glycerol, which significantly stabilize proteins in bulk solution, do not measurably affect their stability when they are attached to a hydroxyl-terminated surface. In contrast, we recovered bulk solution-like stabilization when the attachment surface was instead carboxyl-terminated. It thus appears that chemistry-specific surface interactions can dramatically alter the way in which biomolecules interact with other components of the system.