Asymmetric distribution of cooperativity in the binding cascade of normal human hemoglobin. 2. Stepwise cooperative free energy.
Stepwise cooperative free energies and intermediate Hill coefficients are used to assess the presence of noncooperative sequences in the database of binding free energies previously obtained for the eight partially ligated intermediates of human hemoglobin, encompassing a variety of hemesite analog substitutions. This analysis is prompted by the observed noncooperative binding of two ligands to hemoglobins that are partially substituted with Zn2+-heme, an analog of deoxy Fe2+-heme (Holt et al. (2005) Biochemistry 44, XXXXX). The results show that noncooperative binding sequences are observed in all hemesite analog studied to date. The noncooperative binding observed in (alpha2Znbeta2FeO2) and (alpha2FeO2beta2Zn) is therefore not a Zn-specific substitution artifact. One of several binding sequences from singly to triply ligated hemoglobin is also observed to occur with little or no positive cooperativity. These results demonstrate the variability possible among different ligation pathways in a highly cooperative multi-subunit system such as hemoglobin. As a direct consequence of this variability, differences among ligation pathways are not always detectable using cooperativity functions based on statistical distributions, such as the Hill coefficient n(H). The limitations of Hill coefficient analysis in evaluating cooperativity in intermediates of complex systems is contrasted with the utility of the stepwise binding parameters.
Holt
Jo M
JM
Department of Biochemistry & Molecular Biophysics, Washington University School of Medicine, 660 South Euclid Avenue Box 8231, St. Louis, Missouri 63110, USA.
Ackers
Gary K
GK
D004735:Energy Transfer; D006454:Hemoglobins / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D010100:Oxygen / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D013816:Thermodynamics
D006454:Hemoglobins; D010100:Oxygen