COORDINATION-COMPLEXES AND CATALYTIC PROPERTIES OF PROTEINS AND RELATED SUBSTANCES .109. ELECTROSTATIC STABILIZATION IN MYOGLOBIN - PH-DEPENDENCE OF SUMMED ELECTROSTATIC CONTRIBUTIONS

The modified Tanford-Kirkwood theory of Shire et al. [Shire, S. J., Hanania, G. I. H., & Gurd, F. R. N. (1974) Biochemistry 13, 2967] for electrostatic interactions was applied to compute the free energy contributions from individual pairs of charge loci in sperm whale ferrimyoglobin. Such interaction energies depend not only on the fractional occupancy of each charge site but also on their static solvent accessibilities, in addition to the geometrical and other factors inherent to the treatment. The pH-dependent unfolding of the myoglobin in acid solution is strongly influenced by ionic strength in such a way that the native form is relatively destabilized by increased ionic strength. Since the titration behavior of the denatured form is less sensitive to ionic strength variation than that of the native form, it follows that the native form experiences net stabilization from intramolecular@r@nelectrostatic interactions. The unfolded forms are likewise stabilized by ionic equilibria as a result of protonation of histidine residues that are masked in the native state but exposed in the denatured state. The summed electrostatic free energy of the native structure shows a broad maximum at about pH 6.5, in keeping with the observed thermal stability maximum [Acampora, G., & Hermans, J., Jr. (1967) J. Am. Chem. Soc. 89, 1543], with a net maximum stabilization of approximately 10.6 kcal/mol at 0.00 M, 10.0 kcal/mol at 0.01 M, and 8.6 kcal/mol at 0.10 M ionic strength. By difference it may be estimated that 5-6 kcal/mol of stabilization of the native protein structure can be ascribed to nonionic factors. The charge sites play a subtle dual role in both stabilizing and enhancing the water solubility of the protein. © 1979, American Chemical Society. All rights reserved.