Quaternary structure has little influence on spin states in mixed-spin human methemoglobins

Philo, John S.; Dreyer, Ulrich

doi:10.1021/bi00333a027

Cited by 21 publications

(16 citation statements)

References 30 publications

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“…However, they do demonstrate that this is still an open question which warrants further investigation. That this is the case is also indicated by the results of Philo and Dreyer [19]. These authors report only modest values of AAG" for the thiocyanate derivative of human hemoglobin as a result of P6-inositol addition.…”

Section: Aag" = -R T Ln-= R T L N ( P+ -P?s 2 ) X ( P K -$ )mentioning

confidence: 70%

Studies on the linkage between spin equilibria and protein structure in carp ferric hemoglobin

Noble

Young

Vitale

et al. 1987

European Journal of Biochemistry

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The effects of protein conformation on the spin-state equilibria of several derivatives of carp hemoglobin have been examined. This has been done by measuring the pH dependence of the paramagnetic susceptibilities of these derivatives in the presence and absence of inositol hexakisphosphate, P6-inositol. In all cases the addition of P6-inositol at low pH and the lowering of the pH in the presence of P6-inositol shift the spin-state equilibrium in favor of the high-spin electronic configuration. The P6-inositol and pH dependence of these magnetic properties parallels the pH and P6-inositol dependence of the conformational state of the hemoglobin as determined in earlier studies and further supports a thermodynamic linkage between the electronic state of the iron atoms and the quaternary structure of the hemoglobin molecule.When oxygen binds to hemoglobin there is an alteration in the structure of this protein from the low-affinity, deoxy structure to the high-affinity, ligand-saturated structure. This conformational transition appears to be an essential feature of the cooperative binding of oxygen, carbon monoxide and other heme ligands [l, 21. Such cooperativity is observed as an increase in the ligand affinity of the hemoglobin molecule as the four oxygen-binding or ligand-binding sites become progressively saturated. Under normal circumstances this structural transition in human hemoglobin requires the binding or release of ligand. In contrast to this, the hemoglobin of the carp can be switched between a similar pair of structural states by specific binding or removal of organic phosphate or protons, all of which bind at sites other than the heme group [3]. This remarkable property permits one to examine the effects of protein structure on the ligand-binding site without the need to change the level of saturation or the nature of the ligand. This behavior is found for both ferrous and ferric derivatives of carp hemoglobin [3,4].It is well known that for many ferric derivatives of heme proteins the energy splitting between the low-spin, S = 112, and high-spin, S = 512, states of iron is small enough for a thermal equilibrium to exist between these electronic configurations [5, 61. This spin-state equilibrium depends on the nature of the ligand and varies from one protein to another. In the case of hemoglobin it has also been shown to depend on the structure of the protein. quantify this linkage at room temperature in aqueous solution for carp aquomethemoglobin and azide methemoglobin [lo]. This work has now been extended to several other derivatives of carp hemoglobin and to the determination of the pH dependence of susceptibility in the presence and absence of 1 mM inositol hexakisphosphate (P6-inositol). MATERIALS AND METHODSMagnetic susceptibilities were measured with a superconducting fluxmeter which has been previously described 18, 91. Calibration and procedures for changing samples result in measurements with an overall accuracy of 0.03% of the susceptibility of water in the room temperature range. To co...

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Section: Aag" = -R T Ln-= R T L N ( P+ -P?s 2 ) X ( P K -$ )mentioning

confidence: 70%

Studies on the linkage between spin equilibria and protein structure in carp ferric hemoglobin

Noble

Young

Vitale

et al. 1987

European Journal of Biochemistry

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“…In contrast, the smaller effect of bezafibrate alone continues to increase as the pH is lowered. Philo and Dreyer (1985) reported that the nitrite derivative of human hemoglobin was unstable in their hands. To control for the possibility of time-dependent changes, spectra of solutions of this derivative were examined as a function of time.…”

Section: Resultsmentioning

confidence: 99%

Spin equilibria in human methemoglobin: effects of bezafibrate and inositol hexaphosphate as measured by susceptometry and visible spectroscopy

Noble¹,

DeYoung²,

Vitale³

et al. 1989

Biochemistry

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The effects of inositol hexaphosphate (IHP) and a second allosteric effector, bezafibrate, on the spin-state equilibria of the mixed-spin derivatives of ferric human hemoglobin A are examined. Changes in spin-state equilibrium are monitored by measuring absorption spectra in the visible region (460-700 nm) as well as by direct measurements of magnetic susceptibility by means of a superconducting fluxmeter. The addition of IHP at pH 6.5 results in a measurable shift in the spin equilibria of these derivatives toward higher spin. However, the addition of bezafibrate in the presence of IHP results in still larger shifts toward the high-spin form. The changes in the free energies of the spin-state equilibria resulting from the combination of these two effectors are similar in magnitude to that which results from the R-state to T-state transition in carp hemoglobin.

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“…When Philo and Dreyer repeated the magnetic experiments with both human HbA and the abnormal human Hb Kansas in which the R-structure is destabilized by the rupture of a crucial hydrogen bond, they found no significant rise in the paramagnetic susceptibility of either Hb on switching their azide-met derivatives from R to T and concluded that this rise was a peculiarity shown only by fish Hbs (40). This seemed improbable in view of the extensive homology of the heme surrounds and subunit contacts in all vertebrate Hbs.…”

Section: Changes In Spin Equilibrium Of the Ironsmentioning

confidence: 99%

The Stereochemical Mechanism of the Cooperative Effects in Hemoglobin Revisited

Perutz

Wilkinson

Paoli

et al. 1998

Annu. Rev. Biophys. Biomol. Struct.

528

524

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In 1970, Perutz tried to put the allosteric mechanism of hemoglobin, proposed by Monod, Wyman and Changeux in 1965, on a stereochemical basis. He interpreted their two-state model in terms of an equilibrium between two alternative structures, a tense one (T) with low oxygen affinity, constrained by salt-bridges between the C-termini of the four subunits, and a relaxed one (R) lacking these bridges. The equilibrium was thought to be governed primarily by the positions of the iron atoms relative to the porphyrin: out-of-plane in five-coordinated, high-spin deoxyhemoglobin, and in-plane in six-coordinated, low-spin oxyhemoglobin. The tension exercised by the salt-bridges in the T-structure was to be transmitted to the heme-linked histidines and to restrain the movement of the iron atoms into the porphyrin plane that is necessary for oxygen binding. At the beta-hemes, the distal valine and histidine block the oxygen-combining site in the T-structure; its tension was thought to strengthen that blockage. Finally, Perutz attributed the linearity of proton release with early oxygen uptake to the sequential rupture of salt-bridges in the T-structure and to the accompanying drop in pKa of the weak bases that form part of them. Almost every feature of this mechanism has been disputed, but evidence that has come to light more than 25 years later now shows it to have been substantially correct. That new evidence is reviewed below.

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Quaternary structure has little influence on spin states in mixed-spin human methemoglobins

Cited by 21 publications

References 30 publications

Studies on the linkage between spin equilibria and protein structure in carp ferric hemoglobin

Studies on the linkage between spin equilibria and protein structure in carp ferric hemoglobin

Spin equilibria in human methemoglobin: effects of bezafibrate and inositol hexaphosphate as measured by susceptometry and visible spectroscopy

The Stereochemical Mechanism of the Cooperative Effects in Hemoglobin Revisited

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