PubMed · 2195851
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W A Eaton, J Hofrichter. 1990. Sickle cell hemoglobin polymerization.. https://doi.org/10.1016/s0065-3233(08)60287-9
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The sickling variant of human hemoglobin, Hb S (beta 6 Glu-->Val), assembles into 14-strand helical fibers composed of seven pairs of double strands. The organization of the helical double strands closely resembles the parallel, half-staggered, linear strand pairs of the crystals of Hb S characterized by Wishner et al. In the crystals, the molecules are arranged such that each possesses a beta 6 Val in contact with a molecule on the opposite strand. In the fibers, the overall hexagonal packing of strands leads to 22 classes of potential contacts between the seven double strands, but the presence of 2-fold helical symmetry reduces these contacts to 11 distinct classes. An analysis of the intermolecular contacts reported by Watowich et al., based on the data of Carragher et al., indicated a loosely packed structure for which only four of the 11 potential classes of contacts between double strands are significant (residues within 5 A). We have recently analyzed the packing based on the results of Dykes et al. and Rodgers et al., and compared the findings with the structure derived from the data of Carragher et al. We find serious differences between the two data sets concerning the packing of double strands. The Dykes-Rodgers data indicate a more closely packed structure in which nine of the 11 potential classes of contacts are within 5 A. Considerations on the stability of certain contacts derived from incomplete fibers, as well as studies of Hb molecules composed of beta S chains and mutant alpha chains, suggest that the structural model with closer packing of the double strands provides a better correlation with the other experimental results.
Polymerization of half-liganded Hb S was investigated using Ni(II)-Fe(II) hybrid Hb S, in which heme in either alpha or beta s subunits is replaced by Ni (II) protoporphyrin IX. Studies on the polymerization of these hybrid hemoglobins were carried out under aerobic conditions. Both alpha 2 (Ni) beta 2s (Fe-CO) and alpha 2 (Fe-CO) beta 2s (Ni) polymerized with a distinct delay time as do native deoxy-Hb S and Ni(II) Hb S. However, the critical concentration for polymerization of half-liganded Hb S, alpha 2 (Ni) beta 2s (Fe-CO) and alpha 2 (Fe-CO) beta 2s (Ni), was 4- and 8-times higher, respectively, than that of Ni(II)-Hb S. Kinetics of polymerization of both deoxygenated hybrid hemoglobins with CO completely removed were the same, although the critical concentrations for polymerization were intermediate between those for deoxy-Hb S and Ni(II)-Hb S. These results suggest that the small tertiary conformational change associated with the doubly liganded state may be much less favorable to polymerization than the completely unliganded state of Hb S. The conformational change depends on whether alpha or beta chain is liganded. The ease of polymerization and low solubility of sickle hemoglobin is dependent not only on quaternary, but on tertiary structural changes, as well as on the substitution of Val for Glu at the beta 6 position.
Electron microscopy of sickle cell hemoglobin fibers fixed at different times during gelation shows an exponential distribution of fiber lengths, with many short fibers and few long ones. The distribution does not change significantly with time as polymerization progresses. If this distribution of lengths reflects kinetic mechanism of fiber assembly, it complements information from studies of the progress of average properties of the polymers and, as has been done for other rod-like polymerizing systems, permits testing of models for the mechanism of fiber assembly. In this case, the results are consistent with the double nucleation model of Ferrone et al. or with a related alternative model based on fiber breakage. However, other possible causes of this microheterogeneity exist, including: breakage due to solution shearing of the long, rod-like, fibers; the presence of residual nuclei; equilibrium relations governing polymerization; and breakage of solid-like but weak gels that develop early and adhere to the grid. The arguments against the first three of these possibilities suggest that they are not responsible. However, breakage of entanglements or cross-links in a solid-like and adherent gel is consistent with the distributions.