super(14)N NMR relaxation times of several protein amino acids in aqueous solution - comparison with super(17)O NMR data and estimation of the relative hydration numbers in the cationic and zwitterionic forms

The super(14)N nuclear magnetic resonance (NMR) linewidths of the alpha -amino groups of several protein amino acids were measured in aqueous solution, with and without composite proton decoupling, to estimate the effect of proton exchange and molecular weight on the linewidths. It is shown that, contrary to earlier claims, the increase in the linewidth at low pH is not exclusively due to the effect of proton exchange broadening. The super(14)N linewidths, under composite proton decoupling, increase with the bulk of the amino acid, and increase at low pH. Statistical treatment of the experimental super(14)N and literature super(17)O NMR data was performed assuming two models: (i) an isotropic molecular reorientation of a rigid sphere in a medium of viscosity eta , (ii) a stochastic diffusion of the amino and carboxyl groups comprising contributions from internal ( tau sub(int)) and overall ( tau sub(mol)) motions. Assuming a single correlation time from overall molecular reorientation ( tau sub(mol)), then, a linear correlation was found between the linewidths and the molecular weights of the protein amino acids at the pH values 0.5 and 6.0, which are characteristic of the cationic and zwitterionic forms, respectively. The slopes of the straight-lines were found to be dependent of pH for super(14)N, contrary to the super(17)O linear correlations whose slopes were found to be independent of pH. Assuming effective correlation times of the amino and carboxyl groups, which comprise contributions from the internal ( tau sub(int)) and overall ( tau sub(mol)) motions, then, a significant improvement of the statistics of the regression analysis was observed. The super(14)N relaxation data, in conjunction with super(17)O NMR linewidths, can be interpreted by assuming that the super(14)N quadrupole coupling constants (NQCCs) are influenced by the protonation state of the carboxyl group, the super(17)O NQCCs remain constant, and the cationic form of the amino acids is hydrated by an excess of 1-3 molecules of water relative to the zwitterionic state.