TEMPERATURE-DEPENDENCE OF H-1 CHEMICAL-SHIFTS IN PROTEINS

Temperature coefficients have been measured by 2D NMR methods for the amide and (CH)-H-alpha proton chemical shifts in two globular proteins , bovine pancreatic trypsin inhibitor and hen egg-white lysozyme. The temperature-dependent changes in chemical shift are generally linear u p to about 15 degrees below the global denaturation temperature, and t he derived coefficients span a range of roughly -16 to +2 ppb/K for am ide protons and -4 to +3 ppb/K for (CH)-H-alpha. The temperature coeff icients can be rationalized by the assumption that heating causes incr eases in thermal motion in the protein. Precise calculations of temper ature coefficients derived from protein coordinates are not possible, since chemical shifts are sensitive to small changes in atomic coordin ates. Amide temperature coefficients correlate well with the location of hydrogen bonds as determined by crystallography. It is concluded th at a combined use of both temperature coefficients and exchange rates produces a far more reliable indicator of hydrogen bonding than either alone. If an amide proton exchanges slowly and has a temperature coef ficient more positive than -4.5 ppb/K, it is hydrogen bonded, while if it exchanges rapidly and has a temperature coefficient more negative than -4.5 ppb/K, it is not hydrogen bonded. The previously observed un reliability of temperature coefficients as measures of hydrogen bondin g in peptides may arise from losses of peptide secondary structure on heating.