Enthalpic and entropic components of cooperativity for the partially ligated intermediates of hemoglobin support a "symmetry rule" mechanism.
Subunit assembly reactions of hemoglobin's 10 ligation microstates have been studied as a function of temperature to evaluate the enthalpic and entropic components of cooperativity. It is found that the cooperative enthalpies and cooperative entropies distribute in close agreement with predictions of the symmetry rule mechanism (Ackers et al., 1992) previously deduced from the free energy distribution, in combination with structure-sensitive probes (Doyle & Ackers, 1992; LiCata et al., 1993; Daugherty et al., 1994). Principal findings of the present study are as follows: (1) In unligated hemoglobin (quaternary T), dimer-tetramer assembly is driven by a large negative enthalpy, whereas in the fully ligated (quaternary R) species, the driving force for quaternary assembly is entropic. For the eight intermediate ligation species, the switchover from enthalpic to entropic control follows precisely the symmetry rule predictions; i.e., switching from enthalpic to entropic control occurs at each of the six steps that create ligated heme sites on both sides of the dimer-dimer interface. The combinatorial distribution found previously with free energies does not therefore arise from coincidental enthalpy-entropy compensation that masks a more fundamental distribution. (2) The free energy of tertiary constraint delta Gtc, which pays for intradimer cooperativity prior to quaternary switching, contains large enthalpic and entropic components delta Htc and delta Stc. Like delta Gtc, these terms vanish at the second binding step within the T tetramer. It is found that delta Gtc arises from a net enthalpic dominance over an almost equally large T delta Stc. (3) The stepwise enthalpies correlate with stepwise values of Bohr protons and Bohr free energies (Daugherty et al., 1994) throughout the cascade of 16 stepwise reactions; the correlated clusters of these values follow predictions of the symmetry rule mechanism. (4) These results obtained with cyanomethemoglobin are consistent with the corresponding data on oxygenated hemoglobin which has been resolved at each stage of oxygenation.
Huang
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Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Ackers
G K
GK
D000494:Allosteric Regulation; D006454:Hemoglobins / Q000737:chemistry*; D046911:Macromolecular Substances; D008706:Methemoglobin / Q000031:analogs & derivatives / Q000737:chemistry; D011487:Protein Conformation; D013696:Temperature; D013816:Thermodynamics
D006454:Hemoglobins; D046911:Macromolecular Substances; C007169:cyanomethemoglobin; D008706:Methemoglobin