Determination of the rate constants k1 and k2 of the Linderström-Lang model for protein amide hydrogen exchange. A study of the individual amides in hen egg-white lysozyme.

The pH dependence of the amide/solvent hydrogen exchange of individual amide groups in hen egg-white lysozyme has been studied by nuclear magnetic resonance spectroscopy. Lysozyme has been used here as a model for a globular protein to re-examine the hypothesis for the amide/solvent hydrogen exchange reaction proposed by K. Linderstrøm-Lang and described in detail by Hvidt and Nielsen. The work has been focused on the most slowly exchanging amide at the temperature of 21 degrees C and in the pH range between 4 and 8. Exchange rates have been measured for 64 of the 126 amide protons and the pH dependence has been determined for 52 of these. The amides examined represent a sample that includes all the types of secondary structure and they are placed in the globular structure in a range of 3.2 A to 8.5 A from the closest water molecule on the surface. The measured exchange rates at pH 6 have been compared to these structural parameters and the results suggest that the rate constants are determined partly by the distance to the surface and partly by the type of secondary structure the amide is engaged in. Near the surface and in the very interior the distance to the surface seems to be rate-determining. Between the extremes the type of secondary structure is rate determining. The pH dependent exchange of the examined amides was shown to be in agreement with the Linderstrøm-Lang model. For each of the amides examined the rate constants for the opening and the closing reaction in the first reaction step of the Linderstrøm-Lang model has been calculated and compared to structural parameters.