Light-scattering investigations of the subunit dissociation of human hemoglobin A. Effects of the aliphatic acid salts.

The subunit dissociation of human hemoglobin A by the aliphatic acid salts at neutral pH has been investigated by light-scattering molecular-weight measurements at 630 nm. Dissociation of hemoglobin tetramers to alphabeta dimers is observed in essentially all experiments at low to intermediate levels of salt concentrations, below the denaturation transitions, described in the accompanying paper (Ibanez, V.S., and Herskovits, T.T. (1976), Biochemistry 15, preceding paper in this issue). The effectiveness of the salts as subunit dissociating agent, reflected by the slopes, s, of the plots of deltaGDdegrees, the standard free energy of dissociation, vs. [D], the salt concentration, is found to increase with increasing alkyl chain length or hydrocarbon content of the salt. Estimates of the apparent number of amino acid sites at the areas of contact per alphabeta dimer formed, N', based on the slopes of the higher members of the series have been obtained using the equation, deltaGDdegrees = deltaGD,Wdegrees - 2N'RTKB[D]. Independent estimates of the binding constant, KB, required for these calculations were based on free-energy transfer data of hydrophobic amino acid alkyl groups and protein denaturation data. Our estimates of N' denaturation data. Our estimates of N' obtained with the more reliable data of the higher members of salt series are in the ranges of 19 and 27 amino acid groups, shown by the x-ray crystallographic structure of horse and human hemoglobin of Perutz (Perutz, M.F., et al. (1968), Nature (London) 219, 131) and Fermi ((1975) J. Mol. Biol. 97, 237) for the smaller alpha1 beta2 contact areas in the tetrameric structure. The lower estimates than 27 based on our dissociation of human hemoglobin suggest that several of the amino acid residues in the contact areas of the subunits are partially exposed to solvent. The increasing effectiveness of the higher mn imporant source of stabilization of the tetrameric structure of hemoglobin.