Heats of carbon monoxide binding by hemoglobin M Iwate

Abstract

The heat of reaction of CO gas with the alpha2Mmetbeta2 and alpha2Mbeta2 species of the alpha-chain mutant hemoglobin M Iwate has been studied in buffers with different heats of ionization of 25degrees and in the absence of organic phosphates. For the alpha2Mmetbeta2deoxy species we find a small Bohr effect (0.12 mol of H+/mol of CO) which is in correspondence with that found in equilibrium studies. The heat of reaction, when corrected for proton reaction with buffer, is -18.4 +/- 0.3 kcal/mol of CO at pH 7.4 At pH 9 the same value is observed within experimental error. This value compares closely with heats of reaction of CO with myoglobin and with van't Hoff determinations of the heat of oxygen binding to isolated hemoglobin alpha and beta chains after correction for the heat of replacement of O2 by CO. Furthermore, an analysis of the differential heat of ligand binding as a function of the extent of reaction indicated that, within experimental error, the heat of reaction with the first beta-chain heme in alpha2Mmetbeta2deoxy is the same as the second. Since the quaternary Tleads to R transition is blocked in this mutant hemoglobin, we compared it with Hb A to estimate the enthalpic component of the allosteric T leads to R transition in Hb A. The heats of reaction with CO(g) and Hb A are -15.7 +/- 0.5 and -20.9 +/- 0.5 kcal/mol at pH 7.4 and 9.0, respectively. In going from the T to the R state we find an enthalpy of transition of 9 +/- 2.5 kcal at pH 7.4 and -12 +/- 2.5 kcal at pH 9.0. From published free energies of transsition we conclude the T leads to R transition is enthalpically controlled at p/ 7.4 but entropically controlled at pH 9.0 A near normal Bohr effect is estimated from heats of reaction of CO with alpha2Mdeoxybeta2deoxy in various buffers. A large than normal heat of reaction (-21.6 +/- 0.5 kcal/mol of CO) is attributed to the abnormal alpha chains in Hb M Iwate.

A Calorimetric Study of the CO Bohr Effect of Monomeric Haemoglobins

“…This agrees with their similar equilibrium binding properties [4]. The magnitude of the observed intrinsic heat of CO(g) binding (-19.7 kcal/mol or -82.4 kJ/mol CO) is in the same range observed for inyoglobin from horse (-18 kcal/mol or -75.3 kJ/ mol CO) [9] and a and chains from Hb A, estimated [8] [8]. In the case of Hb A this heat includes the heat of the quaternary T -+ R transition which is exothermic at this pH where the Bohr effect is absent [8].…”

“…1 and 2 we have plotted at pH 7.1 values of -A H V~Y S U S AHbuffcr using values of -0.8 (-3.4), 6.7 (28.0) and 11.2 (46.9) kcal/mol H + (kJ/mol) for the heats of ionization at 25 "C for maleate, bis-Tris and Tris buffers [8,10]. From Eqn (2) The calculated values for AHBohr are 5.7 I- 1.7 and 7.9 f 0.9 kcal/mol (23.8 f 7.1 and 33.2 f 3.8 kJ/mol) for Chironomus haemoglobin I11 and IV, respectively.…”

“…The heat measurements were made at constant volume and corrected by the RT factor to give enthalpy changes, AH, according to Gaud et al [8].…”

Energetics of subunit assembly and ligand binding in human hemoglobin

“…Characterization of ligand binding properties for the subunits in various stages of assembly, and of the assembly reactions themselves, provides an experimental basis for understanding how subunit interaction brings about the observed alterations in affinity and regulatory properties. For normal human hemoglobin a comprehensive set of thermodynamic properties pertaining to these processes has recently been developed (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16). The resulting energetic picture imposes constraints that must be satisfied by any theory proposed to explain the mechanism of cooperative oxygen binding.…”

“…THERMODYNAMIC RESOLUTION OF THE LIGAND-LINKED ASSEMBLY FOR HUMAN HEMOGLOBIN A series of recent experimental studies on human hemoglobin (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16) has provided an extensive and self-consistent set of thermodynamic properties pertaining to (a) the linkage between dimer tetramer association and oxygen binding, (b) the effects of pH and chloride on the oxygenation linked dimer-tetramer reactions, (c) the properties of isolated chains, including their ligand binding and self-association reactions, and (d) the reconstitution of functional hemoglobin tetramers from the constituent subunits. The results summarized here will focus mainly upon the processes of categories a and b.…”