Proton and phosphorus-31 nuclear magnetic resonance investigation of the interaction between 2,3-diphosphoglycerate and human normal adult hemoglobin

Russu, Irina M.; Wu, Shing Shing; Bupp, Keith A.; Ho, Nancy T.; Ho, Chien

doi:10.1021/bi00467a027

Cited by 24 publications

(23 citation statements)

References 44 publications

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“…In vivo, the deoxy T-state is stabilized by salt links mediated by chloride anions at the a N terminus (Chiancone et al, 1975;Arnone & Williams, 1977;Perutz et al, 1994) and diphosphoglycerate (DPG) at the b C terminus (Benesch et al, 1969;Arnone, 1972;Russu et al, 1990). Our usual experimental solutions include pyrophosphate (PPi; 0.02 M) which serves as an analog of DPG.…”

Section: Allosteric Effectors: Phosphate and Chloridementioning

confidence: 99%

Energetic components of the allosteric machinery in hemoglobin measured by hydrogen exchange

Englander

Louie

McKinnie³

et al. 1998

Journal of Molecular Biology

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Section: Allosteric Effectors: Phosphate and Chloridementioning

confidence: 99%

Energetic components of the allosteric machinery in hemoglobin measured by hydrogen exchange

Englander

Louie

McKinnie³

et al. 1998

Journal of Molecular Biology

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“…However, the site of Gra(2,3)P2 binding to oxy-Hb is still uncertain, but two possibilities appear to exist; Gra(2,3)P2 may interact with positively charged residues at the entrance to the central cavity or another site or sites may exist. There is evidence that the binding site is close to that of deoxy-Hb (Gupta et al, 1979;Russu et al, 1990). Given the similar pH dependence of the binding of Gra(2,3)P2 and ATP as well as other anions (Garby et al, 1969;Janig et al, 1970;Rapoport et al, 1972) and the fact that Gra(2,3)P2 decreases the binding of other metabolites and vice versa (Garby et al, 1969;Berger et al, 1973;Hamasaki and Rose, 1974), it is assumed that ATP and other metabolites have the same binding site on both oxy-Hb and deoxy-Hb as does Gra(2,3)P2.…”

Section: Association Associationmentioning

confidence: 99%

Model of the pH‐Dependence of the Concentrations of Complexes Involving Metabolites, Haemoglobin and Magnesium Ions in the Human Erythrocyte

Mulquiney

Kuchel

1997

European Journal of Biochemistry

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The rate of glucose consumption and the concentrations of glycolytic intermediates in human erythrocytes have long been known to be pH sensitive. Despite the extensive literature on modelling erythrocyte metabolism, no model developed so far can adequately describe all of these pH-dependent changes. None of these models have included all the significant association reactions between metabolites, Hb and Mg2+ that will influence metabolism. As part of a larger enterprise to develop a detailed model of erythrocyte glycolysis, we present a sub-model which predicts, as a function of pH and oxygenation state, the concentrations of free and Mg2+-bound metabolites that are substrates, co-factors and effectors of glycolysis. This model shows that pH changes around physiological values can cause large changes in the distribution of metabolites between free, bound and Mg2+-complexed forms, based on binding interactions alone ; in oxygenated cells, at pH 7.2 -7.6, many glycolytic intermediates undergo changes in concentration of 50-100 %. The model also predicts intracellular concentrations of free Mg2+ in erythrocytes to be 0.4 mM and 0.64 mM in oxygenated and deoxygenated cells, respectively, assuming a total magnesium concentration of 3 mM (~8 8 % of the total magnesium usually found in erythrocytes). This is in close agreement with the values found by Flatman [Flatman, P. W. (1980) J. Physiol. 300,[19][20][21][22][23][24][25][26][27][28][29][30] Hexokinase has a high 'flux control coefficient' in human erythrocyte glycolysis, so the dependence of its rate on the pH and oxygenation state of haemoglobin is important. With a low oxygen tension and an intracellular pH of 7.34, the major inhibitor of its activity (2,3-bisphosphoglycerate) is 85 % bound to either haemoglobin or MgZf, and the maximum possible flux of substrate via it would be 2.05 mmol L erythrocytes-' h-'. However, if the haemoglobin were saturated with oxygen, and the pH were 7.2, it was calculated that the maximum rate would be 1.48 mmol L erythrocytes-' h-'; this is primarily due to a doubling of the free 2,3-bisphosphoglycerate concentration. However, the full extent of the inhibition is counteracted because the concentration of the Mg2+-2,3-bisphosphoglycerate would be approximately doubled. Many other similar comparisons are possible with this new model, which highlights the complex network of interactions between haemoglobin, Mg", H' and the metabolites as substrates and effectors of the glycolytic reactions.Keywords: erythrocyte glycolysis ; regulation of metabolism ; pH effect; magnesium ions ; computer simulation.Many metabolically significant ions, such as ATP, 2,3-bisphosphoglycerate [Gra(2,3)P2] and other phosphorylated metabolites in human erythrocytes, bind reversibly with haemoglobin (Hb) and Mg2+. These interactions significantly affect the free and bound concentrations of metabolites within the cells and thus significantly affect the activity of many enzymes (Gupta et al., 1978b;Hamasaki and Rose, 1974;Gerber et al., 1973;Bunn et al., 1971)...

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“…2,3-diphosphoglycerate (2,3-DPG) and myo-inositol hexakisphosphate (IHP)), protons, and chloride ions (1)(2)(3)(4)(5), which bind at heterotropic interaction sites, topologically distinct from the heme at which homotropic ligands bind.…”

mentioning

confidence: 99%

Thermodynamics of Inositol Hexakisphosphate Interaction with Human Oxyhemoglobin

Messana

Angeletti

Castagnola

et al. 1998

Journal of Biological Chemistry

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, which are located in the ␤ 1 ␤ 2 dyad axis, where they have been also proposed to interact with 2,3-diphosphoglycerate, whereas the third group does not appear easily identifiable. Calorimetric measurements of the heat associated with IHP binding at different pH values over the same range indicate that IHP binding is mostly enthalpy-driven at pH < 7 and mostly entropydriven at pH > 7.Human hemoglobin (Hb) 1 is functionally modulated by several non-heme ligands, such as organic phosphates (i.e. 2,3-diphosphoglycerate (2,3-DPG) and myo-inositol hexakisphosphate (IHP)), protons, and chloride ions (1-5), which bind at heterotropic interaction sites, topologically distinct from the heme at which homotropic ligands bind.The structure of this binding pocket has been determined for the interaction of 2,3-DPG (6), which has been shown to bind at the interface between the two ␤-chains, mainly involving three residues from either one of ␤-subunits (i.e. HisNA2(␤2), Lys-EF6(␤82), and HisH21(␤143), see Ref. 6).In more recent years, another organic phosphate, namely IHP (closely related to the inositol pentaphosphate, which is the physiological effector in avian erythrocytes; see Ref. 7), has often been employed to study the modulation of functional properties of human Hb (8, 9). It possesses additional negative charges with respect to 2,3-DPG, and it displays a much larger effect, which suggests the occurrence of additional electrostatic interactions with respect to 2,3-DPG, as from early model building studies on deoxy Hb (10). Therefore, the enhanced functional effect of IHP on the O 2 binding properties of human Hb with respect to 2,3-DPG could be related to a more widespread interaction surface, with the possibility of modulating ligand-linked conformational changes taking place over a larger portion of the whole tetramer.However, a comprehension of the origin for this enhanced effect starts from the characterization of the IHP interaction energy with deoxyHb and with oxyHb. Previous studies have shown that IHP binds HbO 2 , and its binding properties are pH-dependent (11,12). In this study, we have carried out a detailed analysis of the interaction of IHP with human HbO 2 , measuring the effect on (a) proton titration, (b) O 2 dissociation kinetics from fully liganded tetramer, and (c) heat associated to the reaction in order to give a quantitative description of the system and of the interplay between IHP and proton interaction with human HbO 2 . EXPERIMENTAL PROCEDURESHuman HbO 2 was obtained from the blood of healthy volunteers and stripped of anions according to the procedure reported by Riggs (13). Cells were washed three times with iso-osmotic NaCl solutions by centrifugation at 1000 ϫ g, and packed cells were lysed by adding 2 volumes of cold bidistilled water. Stroma were removed by centrifugation at 12,000 ϫ g for 30 min. Hemolysate was first filtered through a Sephadex G-25 column, equilibrated with 0.01 M Tris/HCl buffer, pH 8.0, and EDTA 10 Ϫ5 M, and afterward it was passed through a column of mixed bed ...

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Proton and phosphorus-31 nuclear magnetic resonance investigation of the interaction between 2,3-diphosphoglycerate and human normal adult hemoglobin

Cited by 24 publications

References 44 publications

Energetic components of the allosteric machinery in hemoglobin measured by hydrogen exchange

Energetic components of the allosteric machinery in hemoglobin measured by hydrogen exchange

Model of the pH‐Dependence of the Concentrations of Complexes Involving Metabolites, Haemoglobin and Magnesium Ions in the Human Erythrocyte

Thermodynamics of Inositol Hexakisphosphate Interaction with Human Oxyhemoglobin

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