In situ studies of protein conformation in supercritical fluids: trypsin in carbon dioxide

The conformation of the monomeric enzyme trypsin has been studied in supercritical carbon dioxide. Steady-state fluorescence spectroscopy is used to follow the conformation of trypsin in situ as a function of CO2 density. Our results show for the first time that protein denaturation can occur during the fluid compression step and that the native trypsin is only slightly more stable (1.2 kcal/mol) than the unfolded form. These results demonstrate the power of fluorescence spectroscopy as a tool for studying protein conformation and dynamics in supercritical fluids.