Interaction of Butene with Human Hemoglobin A

Poyart, Claire; Bursaux, E; Guesnon, P; Bohn, B.

doi:10.1111/j.1432-1033.1979.tb13234.x

Cited by 10 publications

(5 citation statements)

References 8 publications

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“…The solubility and kinetics of polymerization of the mutated Hb were compared with those of native Hb S. Under these conditions and after a characteristic lag period, the assembly of Hb S into fibers resulted in a cooperative increase in turbidity at 700 nm until a plateau was achieved. The Hb solubility (C sat ) was determined by measuring the Hb concentration of the soluble phase after completion of the polymerization process (20,21).…”

Section: Methodsmentioning

confidence: 99%

Influence of the A Helix Structure on the Polymerization of Hemoglobin S

Lesecq

Baudin

Kister

et al. 1997

Journal of Biological Chemistry

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Hb S variants containing Lys-␤132 3 Ala or Asn substitutions were engineered to evaluate the consequences of the A helix destabilization in the polymerization process. Previous studies suggested that the loss of the Glu-␤7-Lys-␤132 salt bridge in the recombinant Hb ␤E6V/E7A could be responsible for the destabilization of the A helix. The recombinant Hb (rHb) S/␤132 variants polymerized with an increased delay time as well as decreased maximum absorbance and Hb solubility values similar to that of Hb S. These data indicate that the strength of the donor-acceptor site interaction may be reduced due to an altered conformation of the A helix. The question arises whether this alteration leads to a true inhibition of the polymerization process or to qualitatively different polymers. The oxygen affinity of the ␤132 mutated rHbs was similar to that of Hb A and S, whereas the cooperativity and effects of organic phosphates were reduced. This could be attributed to modifications in the central cavity due to loss of the positively charged lysine. Since Lys-␤132 is involved in the stabilization of the ␣1-␤1 interface, the loss of the ␤132(H10)-␤128(H6) salt bridge may be responsible for the marked thermal instability of the ␤132 mutated rHbs.

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Section: Methodsmentioning

confidence: 99%

Influence of the A Helix Structure on the Polymerization of Hemoglobin S

Lesecq

Baudin

Kister

et al. 1997

Journal of Biological Chemistry

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“…The solubility and the kinetics of polymerization of the mutated Hb were compared to those of native Hb S. Under these conditions, the assembly of Hb S into fibers results, after a characteristic lag period, in a cooperative increase in turbidity (measured by the absorbance at 700 nm) until a plateau was achieved. The Hb solubility (C sat ) was determined by measuring the concentration of the soluble phase after completion of the polymerization process (16,17).…”

Section: Methodsmentioning

confidence: 99%

Functional Studies and Polymerization of Recombinant Hemoglobin Glu-α2β26(A3) → Val/Glu-7(A4) → Ala

Lesecq

Baudin

Kister

et al. 1996

Journal of Biological Chemistry

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In hemoglobin (Hb) S the hydrophobic mutated residue Val-␤6(A3) (donor site) closely interacts with the hydrophobic side groups of Phe-␤85(F1) and Leu-␤88(F4) (EF pocket, acceptor site) of a neighboring tetramer, resulting in decreased solubility and polymerization of the deoxy-Hb. The ␤6(A3) residue is followed by two charged residues Glu-␤7(A4) and Lys-␤8(A5). This cluster has no attraction for the hydrophobic EF pocket. We have modified the ␤7(A4) residue next to the donor site Val-␤6(A3), replacing the charged Glu by a hydrophobic Ala-(rHb ␤E6V/E7A). The single mutant Glu-␤7 3 Ala-(rHb ␤E7A) was also engineered. Both rHbs exhibit a heat instability and an increased oxygen affinity compared to Hb A and Hb S. There was a concentration dependence of the ligand binding properties (1-300 M in heme) indicating an increased amount of dimers relative to Hb A. The deoxy form of rHb ␤E6V/E7A polymerizes in vitro, with a decreased rate of polymer formation relative to Hb S, while the single mutant ␤E7A does not polymerize in the same experimental conditions. The Glu-␤7(A4) 3 Ala substitution does not increase the hydrophobic interaction between donor and acceptor site. We speculate that the loss of the normal saline bridge between Glu-␤7(A4) and Lys-␤132(H10) leads to an increased flexibility of the A helix and may account for the difference of the polymerization for this Hb S mutant.

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“…There was less than 3% difference in total Hb concentration between measurements before and after the polymerization process, indicating that complete solubilization ofthe polymers without denaturation ofthe Hb had been achieved. Parameters of the polymerization [delay time T, total absorbance change (AA), and polymer formation rate k (AA/final time -7)] were computed from the recording traces (3,20,21). Computer modeling of the donor-acceptor site was performed by using the BRUGEL package installed on a VAX 8700 computer (22).…”

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confidence: 99%

Enhanced polymerization of recombinant human deoxyhemoglobin beta 6 Glu----Ile.

Baudin-Chich

Pagnier

Marden

et al. 1990

Proc. Natl. Acad. Sci. U.S.A.

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Polymerization ofthe deoxy form ofsickle cell hemoglobin (Hb S; 136 Glu-*Val) involves both hydrophobic and electrostatic intermolecular contacts. These interactions drive the mutated molecules into long fibrous rods composed of seven pairs of strands. X-ray crystallography of Hb S and electron microscopy image reconstruction of the fibers have revealed the remarkable complementarity between one of the 186 valines ofeach molecule (the donor site) and an acceptor site at the EF corner of a neighboring tetramer. This interaction constitutes the major lateral contact between the two strands in a pair. To estimate the relative importance of this key hydrophobic contact in polymer formation we have generated a modeling of the donor-acceptor interaction shows that the presence of an isoleucine side chain at the donor site induces increased contacts with the receptor site and an increased buried surface area, in agreement with the higher hydrophobicity of the isoleucine residue. The agreement between the predicted and experimental differences in solubility suggests that the transfer of the 136 valine or isoleucine side chain from water to a hydrophobic environment is sufficient to explain the observations.Sickle cell hemoglobin (Hb S) has been the subject of intense interest because the replacement of the charged glutamate residue at the sixth position (A3) in the 13 chains by an uncharged valine leads to a drastic decrease in the solubility of the mutant Hb in its deoxy form. This drives the mutated molecules into long fibrous rods, distorting the erythrocytes into their characteristic sickle shape (1, 2). The thermodynamic behavior and kinetics of polymerization have been studied in detail (for recent reviews see refs. 3 and 4). The long rods in sickled cells possess a highly ordered structure involving 14 helical strands of the mutated tetramers. X-ray analyses of the structure of the polymers (5) as well as molecular imaging studies (6-8) have provided a clear understanding of the mechanisms responsible for the formation of the polymers, which include both hydrophobic and electrostatic interactions between the abnormal tetramers.Three other natural mutations at the 136 position have been described: Hb C (Glu--Lys), Hb Machida (Glu-oGln) (9), and Hb G-Makassar (Glu--Ala) (10). None of these Hbs exhibit gelling properties similar to those of Hb S (10, 11). These reports strongly suggested that the localized change in hydrophobicity at the surface of the A helix in the 13 chains of Hb S is a major determinant in producing the oxygenlinked decrease in solubility of deoxy Hb S, leading to the formation of the polymers. Structural analyses of the fibers and computer molecular modeling have revealed the remarkable complementarity between the side chain of the 136 valine of one tetramer with an acceptor site constituted by the 185 phenylalanine and ,B88 leucine side chains of another tetramer, thereby explaining the primary intermolecular interaction in Hb S fibers. The solubility of the Hb S tetramers may be furth...

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Interaction of Butene with Human Hemoglobin A

Cited by 10 publications

References 8 publications

Influence of the A Helix Structure on the Polymerization of Hemoglobin S

Influence of the A Helix Structure on the Polymerization of Hemoglobin S

Functional Studies and Polymerization of Recombinant Hemoglobin Glu-α2β26(A3) → Val/Glu-7(A4) → Ala

Enhanced polymerization of recombinant human deoxyhemoglobin beta 6 Glu----Ile.

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