Structure of Sickled Erythrocytes and of Sickle-Cell Hemoglobin Fibers

Perutz, M. F.; Bertles, John F.; Döbler, Johanna

doi:10.1073/pnas.70.3.718

Cited by 136 publications

(75 citation statements)

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“…Fibers of deoxyHbS analyzed in this study exhibit an outer diameter of 160-170 A, an inner diameter of about 60 A, and an axial disc-repeat distance of 58 A. The analysis of helical parameters indicates that there are 56 discs per turn stacked normal to the helical axis rather than 48 as suggested by Finch et al (1) and that there are consequently 9.3 rather than 8.0 deoxyHbS molecules per one-sixth of a turn along a helical strand of molecules. These differences in dimensions may result from differences in sample preparation and do not necessarily imply a fundamental difference between the structures.…”

Section: Discussionmentioning

confidence: 61%

“…In this communication we report a preliminary characterization by electron microscopy of one type of ordered aggregate of deoxyHbS molecules, the 6-fold symmetric helix similar to that initially described by Finch et al (1). Our electron micrographs also reveal many disc-like structures which appear to be rings of six hemoglobin molecules with apparent 6-fold symmetry and could represent a substructural component of the fibers.…”

mentioning

confidence: 64%

“…The erythrocyte sickling phenomenon of sickle cell anemia is associated with aggregation of hemoglobin S (HbS) molecules into linear arrays or fibers (1)(2)(3)(4)(5)(6)(7)(8)(9). These aggregates form long bundles of fibers within the erythrocyte and are believed to be responsible for deformation of the cell into abnormal shapes with the consequent acute physiologic manifestations known as sickle cell crisis.…”

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confidence: 99%

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Electron microscopy of fibers and discs of hemoglobin S having sixfold symmetry

Ohtsuki

White

Zeitler

et al. 1977

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

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Aggregated forms of deoxyhemoglobin S were examined with a field emission transmission electron microscope. Images of isolated helical fibers were obtained from sickled cell lysates stained directly on the electron microscope grid. Optical and digital analyses of the electron micrographs showed that the fibers are similar to those characterized by J. T. Finch, M. F. Perutz, J. F. Bertles, and J. Dobler [(1973) Proc. NatL Acad. Sci. USA 70, 718-7221 in that they consist of stacked discs each composed of six hemoglobin molecules. The fibers exhibit an outer diameter of 160-170 A and an inner diameter of about 60 A with an axial spacing of 58 A per disc. The fiber can be described as a helix consisting of 56 discs per helical turn. We observed discs of six hemoglobin molecules, which may be stable substructural components of the fibers. They were observed in preparations of hemoglobin fibers and exhibited 6-fold symmetry by power spectrum analysis. A reconstructed image of a disc digitally filtered for 6-fold symmetry has a maximum external diameter of -170 A and a central hole of 60 A diameter and is similar to the axial projection of a single disc from a low-resolution, three-dimensional reconstructed model of a fiber.The erythrocyte sickling phenomenon of sickle cell anemia is associated with aggregation of hemoglobin S (HbS) molecules into linear arrays or fibers (1-9). These aggregates form long bundles of fibers within the erythrocyte and are believed to be responsible for deformation of the cell into abnormal shapes with the consequent acute physiologic manifestations known as sickle cell crisis. The molecular basis of sickle cell anemia is a mutation in the 13-globin gene causing replacement of Glu A3(6)# with valine in each of the subunits of hemoglobin (11).It is clearly desirable to relate the structural alteration of the hemoglobin molecule to the mechanism responsible for the formation of the characteristic linear aggregates. Structural features of aggregates of deoxyhemoglobin S (deoxyHbS) molecules have been characterized by electron microscopy (1-5), x-ray diffraction (6), and physical chemical studies (7)(8)(9)(10). On the basis of these studies, models have been presented that are consistent with the fl6 site at the points of intermolecular contact (12); however, direct structural determination of the molecular orientation of the deoxyHbS molecule in the fiber is required to resolve the role of the substituted valine residue.Correlating the features of the fibers with the chemical and physical properties of the deoxyHbS molecule has been complicated by the fact that at least two distinct types of fibers have been described by electron microscope studies: a six-stranded helix (1, 2) having 6-fold axial symmetry and two types ofThe costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact. eight-stranded helices in which pairs of st...

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Section: Discussionmentioning

confidence: 61%

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confidence: 64%

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confidence: 99%

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Electron microscopy of fibers and discs of hemoglobin S having sixfold symmetry

Ohtsuki

White

Zeitler

et al. 1977

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

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“…The diffraction maximum at (52 A)-' is probably associated with the molecular packing in the polymeric structure and is independent of the packing of polymers into regular arrays. It corresponds to the diameter of hemoglobin along the molecular z axis (2).…”

Section: Resultsmentioning

confidence: 99%

X-ray diffraction studies of fibers and crystals of deoxygenated sickle cell hemoglobin.

Magdoff-Fairchild¹,

Chiu²

1979

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

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Paracrystalline fibers of deoxygenated sickle hemoglobin in erythrocytes or concentrated solutions exhibit a phase transformation to a fully crystalline state. X-ray diffraction patterns of the fiber and crystallites are similar except in two respects: the equatorial spacings of the fibers suggest that they pack into a square lattice with a = 220 A, whereas those of the crystals can be indexed on the basis of a net of 187 A by 54 A, and the second-order near-meridional reflections are strong on the fiber pattern but weak on that of the crystallites. The crystallites are isomorphous with single crystals gown in polyethylene glycol solution at pH 4.5 whose structure has been determined at near-atomic resolution (Wishner, B. C., Ward, K. B., Lattmen, E. E. & Love, W. E. (1975) J. Mol. Biol. 98, 179-194). Double filaments of molecules with an axial repeat of 64 A comprise the basic unit of both the crystal and fiber structures. Each filament of the pair is translated with respect to its neighbor by half a molecular diameter along the fiber axis. The two filaments are held together by contacts made by Val 6ft in the molecules of one strand with hydrophobic side chains of the molecule in the neighboring strand. This interaction is probably the cause of the aggregation of filaments into fibers that leads to the sickling of erythrocytes.X-ray diffraction patterns (1) of polymers found in sickled erythrocytes of individuals homozygous for sickle cell hemoglobin (Hb S) and in concentrated solutions of deoxygenated Hb S show a series of sharp meridional reflections arising from a 64-A repeat along the fiber. Potentially, these diffraction patterns contain information about intermolecular contact regions within the polymer, but, because of the poor resolution inherent in cylindrically averaged fiber diagrams and because insufficient parallel alignment of fibers further decreases the resolution, the surface lattice of the polymer has not as yet been established from these patterns; without it the intermolecular interactions cannot be determined.The packing of molecules into the polymers should be evident from electron micrographs, but these show several polymorphic forms, including a 6-stranded microtubular structure (2) and 14-stranded solid elliptical cylinders (3). Occasional sheetlike structures (4), and six-membered discs (5) have also been observed. Such a wide range of polymorphism is difficult to reconcile with our diffraction patterns, all of which appear similar except for differences in resolution. The patterns most likely arise from a single diffracting system.On re-examination of deoxygenated sickled erythrocytes sealed in capillaries several years ago, we have discovered strongly birefringent ribbonlike structures as well as bundles of needles of about 3-&,m diameter and 1-cm length. These were in equilibrium with a water clear solution, which showed that they could not be fibers because the critical equilibrium concentration of the polymer-monomer system of deoxy-Hb S is about 0.22 g-cm-3 (6) under th...

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“…This shape change results from aggregation of the sickle hemoglobin (HbS) molecules into parallel filaments [1][2][3][4][5]. Studies of this protein aggregation process have provided much useful information about the molecular mechanism of sickling [1,2,4,[6][7][8][9].…”

Section: Introductionmentioning

confidence: 99%