How does a protein unfold on a reversed-phase liquid chromatography surface?

Nuclear magnetic resonance and isotope-exchange techniques were used to stu dy unfolding of lysozyme adsorbed to reversed-phase liquid chromatography s urfaces. All surfaces resulted in significant amide exchange, indicating so lvent exposure and some loss of native structure. However, none of the surf aces resulted in complete exchange. The greatest amount of structure was pr eserved on the C-4 silica, with the most protection in the alpha-helix doma in. C-18 silica and Source RPLC resulted in much greater solvent exposure. No simple correlation was found between chromatographic retention and degre e of surface unfolding. Variations in residual conformation may explain the complex retention behavior of proteins vs. small molecules. (C) 1999 Elsev ier Science B.V. All rights reserved.