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S J Gill

Publications and source records attributed to S J Gill.

13 recordsLinked to original sources
PubMed1992

Oxygen binding constants and stepwise enthalpies for human and bovine hemoglobin at pH 7.6.

A high-precision thin-layer gas-solution microcalorimeter has been developed to study the binding reactions of gaseous ligands with ligand-binding macromolecules in a manner analogous to that of the Gill thin-layer optical apparatus [Doleman & Gill (1976) Anal. Biochem. 87, 127]. We have generated differential heat-binding curves of oxygen binding to human and bovine hemoglobin in phosphate buffer at pH 7.6, with the enzyme-reducing system of Hayashi et al. [(1973) Biochim. Biophys. Acta 310, 309]. Experiments were conducted at a number of different temperatures in order to expand the data field, allowing for separation of enthalpy and free energy parameters. This type of experimental analysis makes no assumptions of optical linearity between the various heme groups and reveals that the triply ligated species is measurably significant for both human and bovine hemoglobin. It was also determined that the concentration of doubly ligated species of bovine hemoglobin is relatively low. The experiments indicate that the reactions for both hemoglobins are enthalpy-driven for oxygen stepwise additions 1, 2, and 4 while being entropy-driven for step 3. Human hemoglobin oxygen-binding experiments were also performed with the Gill thin-layer optical apparatus under solution conditions identical to those used in the calorimeter. The experiments revealed that if optical linearity is assumed, the overall third equilibrium constant is negative or near zero. This indicated that either the optical cell's performance is much poorer than the thin-layer calorimeter or there is an appreciable nonlinear optical effect.

Animals↗
PubMed1979

Thermodynamic analysis of carbon monoxide binding by hemoglobin trout I.

Calorimetric measurements at 25 degrees of the differential heat of CO binding by hemoglobin trout I have been examined together with the CO binding isotherms for the protein at 4 degrees and 20 degrees. Simultaneous treatment of these data sets by a statistically rigorous technique permits evaluation of all the thermodynamic parameters for both the Adair and the Monod, Wyman, Changeux (MWC) models. The results show the details of the unusual temperature dependent cooperativity which this hemoglobin exhibits. In the Adair formalism the increasingly favorable free energy change for successive steps of ligand binding are nearly linearly paralleled by increasingly negative enthalpy changes for these steps. This causes the enhanced cooperativity observed as the temperature is decreased. For the MWC case, lowering the temperature increases the stability of the unligated T state relative to the unligated R state since the enthalpy of the T leads to R transition is 29.4 kcal mol-1. Simultaneously, the favorability of ligating R forms relative to T is enhanced since R form ligation is 14.1 kcal (mol CO)-1 more exothermic than that of T. The balance between these opposing effects is to increase ligand binding cooperativity at low temperatures. The predicted temperature dependence of the Hill coefficient for the MWC and Adair models is identical at low and intermediate temperatures, but, interestingly, would show a strong divergence at high temperatures where negative cooperativity is suggested for the Adair case and positive cooperativity for the MWC case.

Animals↗
PubMed1978

Aggregation effects on oxygen binding of sickle cell hemoglobin.

Deoxygenation of concentrated solutions (0.33 gram per milliliter) of sickle cell hemoglobin show (i) a "crisis point" where the oxygen binding curve is unusually steep (Hill coefficient of 5 to 6), and (ii) a simultaneous increase in light scattering. Nearly identical oxygen binding curves are obtained upon oxygenation and deoxygenation of these solutions. The influence of aggregation is to shift the curve toward higher pressures.

Allosteric Regulation↗
PubMed1978

Analysis of ligand binding curves in terms of species fractions.

The ligand binding curve for a macromolecular system presents the average number of ligand molecules bound per macromolecule as a function of the chemical potential or the logarithm of the ligand concentration. We show that various observable properties of this curve, for example its asymptotes and derivatives, are expressible in terms of linear combinations of the mole fractions alphai of macromolecules binding i molecules of ligand. Whenever enough such properties of the binding curve are known, the linear equations in alphai can be solved to give the mole fractions of each of the various macromolecular species. An application of these results is that a Hill plot for hemoglobin-ligand equilibrium where the asymptotes approach unit slope can be made to yield the four Adair constants by a simple algebraic method. A second use is that a knowledge of the first and second derivatives of the binding curve at points along the curve can yield the species fractions as functions of the degree of saturation without direct knowledge of the ligand binding constants. These methods are illustrated by some numerical examples.

Binding Sites↗
PubMed1976

A calorimetric study of the CO Bohr effect of monomeric haemoglobins.

A calorimetric study has been made of the heats of CO reaction with the monomeric haemoglobins of Chironomus thummi thummi III and IV as a function of pH. The number of Bohr protons released at pH 7.1 was determined from heats of reaction in different buffers as 0.19 and 0.31 mol H+/mol CO for haemoglobin III and IV respectively. The heat of the Bohr ionization process was found to be 6 and 8 kcal/mol H+ (25 and 34 kJ/mol) for the haemoglobins III and IV. These values are consistent with values found for histidine groups. A pH-independent part of the reaction enthalpy was determined as - 19.7 kcal/mol CO (-82.4 kJ/mol). The same reaction with myoglobin is less exothermic. From the combination of deltaG0 and deltaH0 values TdeltaS0 values have been calculated. It was found for both haemoglobins that the entropy of reaction is greater by 2 cal K-1 mol-1 (8.4 JK-1 mol-1) at pH 9.5 as compared to pH 6.0.

Animals↗
PubMed1975

Heats of carbon monoxide binding by hemoglobin M Iwate.

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.

Carbon Monoxide↗