Sickle Cell Hemoglobin Polymers Bind Oxygen Noncooperatively

Sunshine, Helen R.; Hofrichter, James; Ferrone, Frank A.; Eaton, William A.

doi:10.1007/978-1-349-06488-5_41

Cited by 2 publications

(3 citation statements)

References 21 publications

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“…One assay, devised by Benesch et al (18), is based on the sharp decrease in Hb O 2 affinity associated with Hb S polymerization and gelation. In experienced hands, this can be a fairly accurate scanning assay of the minimum Hb concentration for gelation, albeit with some limitations (19,20); but each measurement requires multiple determinations of Hb O 2 affinity (p50) and for micro-scale determinations requires equipment (such as the Hem-O-Scan from Aminco, no longer manufactured) that is not readily available.…”

Section: Discussionmentioning

confidence: 99%

Polymer Structure and Solubility of Deoxyhemoglobin S in the Presence of High Concentrations of Volume-excluding 70-kDa Dextran

Bookchin

Balazs

Wang

et al. 1999

Journal of Biological Chemistry

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Earlier observations indicated that volume exclusion by admixed non-hemoglobin macromolecules lowered the polymer solubility ("C sat ") of deoxyhemoglobin (Hb) S, presumably by increasing its activity. In view of the potential usefulness of these observations for in vitro studies of sickling-related polymerization, we examined the ultrastructure, solubility behavior, and phase distributions of deoxygenated mixtures of Hb S with 70-kDa dextran, a relatively inert, low ionic strength space-filling macromolecule. Increasing admixture of dextran progressively lowered the C sat of deoxyHb S. With 12 g/dl dextran, a 5-fold decrease in apparent C sat ("dextran-C sat ") was obtained together with acceptable sensitivity and proportionality with the standard C sat when assessing the effects of non-S Hb admixtures (A, C, and F) or polymerization inhibitors (alkylureas or phenylalanine). The volume fraction of dextran excluding Hb was 70 -75% of total deoxyHb-dextran (12 g/dl) volumes. Electron microscopy showed polymer fibers and fiber-tocrystal transitions indistinguishable from those formed without dextran. Thus when Hb quantities are limited, as with genetically engineered recombinant Hbs or transgenic sickle mice, the dextran-C sat provides convenient and reliable screening of effects of Hb S modifications on polymerization under near-physiological conditions, avoiding problems of high ionic strength.The solubility of deoxyHb 1 S polymers at equilibrium with the tetrameric "monomers" is a basic thermodynamic property of the polymerization process that underlies red cell sickling. Measurement of the polymer solubility, "C sat "(1) (also termed C s (2) or C sol (3)), of Hb S in conditions similar to those within the cells has served as a critical estimate of the relative polymerization tendency when comparing the native or modified Hb S, alone or in mixtures with non-S Hbs or inhibitors of polymerization. A concentrated solution of Hb S is fully deoxygenated to form a gel, and after high speed centrifugation to sediment the polymerized solid phase, the concentration of soluble Hb in the supernatant is the measured C sat .It is well documented that volume exclusion by macromolecules admixed with proteins results in their increased activity and corresponding interactions (4). Thus, when protein macromolecules capable of interacting, such as deoxyHb S, occupy a substantial fraction of the total volume of their medium, the addition of a second species of noninteracting macromolecules crowds the protein molecules closer together, increasing their interactions. More specifically, addition of macromolecules that do not copolymerize with deoxyHb S reduces its polymer solubility by this mechanism (5, 6). However, for the use of such observations in further investigations on the properties of deoxyHb S polymers, it is first necessary to characterize the morphology and properties of the polymer fibers generated in volume-excluded domains by comparison with those previously characterized for standard conditions.In the course of our ...

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Section: Discussionmentioning

confidence: 99%

Polymer Structure and Solubility of Deoxyhemoglobin S in the Presence of High Concentrations of Volume-excluding 70-kDa Dextran

Bookchin

Balazs

Wang

et al. 1999

Journal of Biological Chemistry

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“…Experimental results suggest that this polymer-associated water is inaccessible to soluble Hb and presumably to other macromolecules [9,13,30,39,40]. But the extent to which small solutes may be globally or selectively excluded from polymer water was unknown, and the information is necessary for the analysis of the osmotic effects of lib S polymerization.…”

Section: Introductionmentioning

confidence: 98%

“…Since the generally accepted current estimate of the concentration of Hb within the polymer, Cp (69 + 6 g/dl), is considerably lower than that of Hb in its exclusion volume (34 g/0.25 dl ~ 136 g/dl), polymers of deoxy-Hb S must retain a substantial proportion of water within their lattice structure [9,13,32,40]. Experimental results suggest that this polymer-associated water is inaccessible to soluble Hb and presumably to other macromolecules [9,13,30,39,40].…”

Section: Introductionmentioning

confidence: 99%

Osmotic effects of protein polymerization: Analysis of volume changes in sickle cell anemia red cells following deoxy-hemoglobin S polymerization

Lew

Bookchin

1991

J. Membrain Biol.

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Polymerization-depolymerization of proteins within cells and subcellular organelles may have powerful osmotic effects. As a model to study these we analyzed the predicted volume changes following hemoglobin (Hb) S polymerization in sickle cell anemia (SS) red cells with different initial volumes. The theoretical analysis predicted that dehydrated SS red cells may sustain large polymerization-induced volume shifts whose direction would depend on whether or not small solutes were excluded from polymer-associated water. Experiments with SS cells from promptly fractionated venous blood showed oxygenation-induced swelling, maximal in the densest cells, in support of nonexclusion models. The predicted extent of cell dehydration on polymerization was strongly influenced by factors such as the dilution of residual soluble Hb and the increased osmotic contribution of Hb in cells dehydrated by salt loss, largely overlooked in the past. The osmotic effects of polymer formation may thus play an important part in microcirculatory infarction by dense SS cells, as they become even denser and stiffer during deoxygenation in the capillaries.

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Sickle Cell Hemoglobin Polymers Bind Oxygen Noncooperatively

Cited by 2 publications

References 21 publications

Polymer Structure and Solubility of Deoxyhemoglobin S in the Presence of High Concentrations of Volume-excluding 70-kDa Dextran

Polymer Structure and Solubility of Deoxyhemoglobin S in the Presence of High Concentrations of Volume-excluding 70-kDa Dextran

Osmotic effects of protein polymerization: Analysis of volume changes in sickle cell anemia red cells following deoxy-hemoglobin S polymerization

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