FRUSTRATED MULTIPLE INTERNAL-REFLECTION INFRARED SPECTROSCOPIC STUDIES ON H-1-H-2 EXCHANGE IN PROTEIN FILMS

Using the technique of frustrated multiple internal reflection infrared spectroscopy on germanium plates the hydrogen-deuterium (1H2H) exchange of protein films was studied. Provided that the films were kept under an atmosphere of at least 90% relative humidity the exchange characteristics were the same as in solutions. A quantitative comparison of the exchange pattern with protein structure data obtained from X-ray spectroscopy and tritium isotope studies proved the reliability of the film technique. As in solution the number of unexchanged hydrogens decreased with decreasing proton concentrations. From this and results on the ionization of outer protein surface carboxylate groups it is concluded that proteins at low pH exist in a compact structure, which is gradually opened with increasing pH by the ionization and subsequent hydration of the carboxyl groups. © 1979.