Molecular Crowding Limits the Role of Fetal Hemoglobin in Therapy for Sickle Cell Disease

Rotter, Maria; Aprelev, Alexey; Adachi, K.; Ferrone, Frank A.

doi:10.1016/j.jmb.2005.02.006

Cited by 28 publications

(30 citation statements)

References 32 publications

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“…Acceleration of slow (transition state limited) protein Large increases in the rate of fiber formation by sickle cell hemoglobin (Rotter et al, 2005), actin associations (Minton, 1983;Minton, 2001a). (Drenckhahn and Pollard, 1986), tubulin (Herzog and Weber, 1978) and fibrin (Wilf et al, 1985).…”

Section: Relevance To Cell Biologymentioning

confidence: 99%

How can biochemical reactions within cells differ from those in test tubes?

Minton

2006

Journal of Cell Science

381

246

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Nonspecific interactions between individual macro-molecules and their immediate surroundings (`background interactions') within a medium as heterogeneous and highly volume occupied as the interior of a living cell can greatly influence the equilibria and rates of reactions in which they participate. Background interactions may be either repulsive, leading to preferential size-and-shape-dependent exclusion from highly volume-occupied elements of volume, or attractive, leading to nonspecific associations or adsorption. Nonspecific interactions with different constituents of the cellular interior lead to three classes of phenomena: macromolecular crowding, confinement and adsorption. Theory and experiment have established that predominantly repulsive background interactions tend to enhance the rate and extent of macromolecular associations in solution, whereas predominately attractive background interactions tend to enhance the tendency of macromolecules to associate on adsorbing surfaces. Greater than order-of-magnitude increases in association rate and equilibrium constants attributable to background interactions have been observed in simulated and actual intracellular environments.

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Section: Relevance To Cell Biologymentioning

confidence: 99%

How can biochemical reactions within cells differ from those in test tubes?

Minton

2006

Journal of Cell Science

381

246

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“…Therefore, Hb F plus such peptides might exhibit anti-sickling effects exceeding those of Hb F alone. This has important implications for gene therapy for sickle cell disease, since anti-polymerization induced by these peptides and Hb F may possibly be accomplished by completely different mechanisms; inhibition by the former via direct interaction with Hb S, and the latter by reduction in Hb S concentration via formation of hybrids which are excluded from Hb S polymers (19). We are screening for such peptides and will evaluate their anti-polymerization properties under near physiological conditions.…”

Section: Discussionmentioning

confidence: 99%

Inhibition of Hemoglobin S Polymerization in Vitro by a Novel 15-mer EF-Helix β73 Histidine-Containing Peptide

et al. 2006

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Our mutational studies on HbS showed that the HbS β73His variant (β6Val and β73His) promoted polymerization, while HbS β73Leu (β6Val and β73Leu) inhibited polymerization. Based on these results, we speculated that EF-helix peptides containing β73His interact with β4Thr in HbS and compete with HbS, resulting in inhibition of HbS polymerization. We, therefore, studied inhibitory effects of 15-, 11-, 7-and 3-mer EF-helix peptides containing β73His on HbS polymerization. The delay time prior to HbS polymerization increased only in the presence of the 15-mer His peptide; the higher the amount, the longer the delay time. DIC image analysis also showed fiber elongation rate for HbS polymers decreased with increasing concentration of the 15-mer His peptide. In contrast, the same 15-mer-peptide containing β73Leu instead of His and peptides shorter than 11 amino acids containing β73His including His alone showed little effect on kinetics of polymerization and elongation of polymers. Analysis by protein-chip arrays showed that only the 15-mer β73His peptide interacted with HbS. CD spectra of the 15-mer β73His peptide did not show a specific helical structure, however, computer docking analysis suggested a lower energy for interaction of HbS with the 15-mer β73His peptide compared to peptides containing other amino acids at this position. These results suggest that the 15-mer β73His peptide interacts with HbS via the β4Thr in the β S -globin chain in HbS. This interaction may influence hydrogen bond interaction between β73Asp and β4Thr in HbS polymers and interfere in hydrophobic interactions of β6 Val leading to inhibition of HbS polymerization.Hb S is a naturally occurring mutation of human tetrameric hemoglobin in which the β subunits have a hydrophobic Val in place of a negatively charged Glu at the β6 position. The consequence of this mutation is that solubility of Hb S decreases when oxy Hb S loses oxygen. When deoxy Hb S becomes oversaturated deoxy Hb S assembles into long, multi-stranded fibers under physiological conditions (1,2). Fiber formation is characterized by a delay time prior to polymerization, which is explained by homogeneous and heterogeneous nucleation. During oversaturated Hb S conditions deoxy Hb S monomers form very small polymers by homogeneous nucleation, and these polymers grow by the end addition of hemoglobin molecules in solution. The surface of these growing fibers also can serve as heterogeneous nucleation sites for further growth of additional polymers (1,3). The polymer then assembles into 14-stranded fibers, which finally form a viscous gel. Intracellular polymers or fibers cause reduction in red blood cell deformability (sickling), leading to obstruction of flow in the microcirculation, thus creating vasooclusion and a wide array of physiological problems including episodes of painful crises (1,4).Computational refinements of x-ray-determined crystal structures clarified the details of many of the axial and lateral contacts in Hb S polymers (5). These results and properties of th...

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“…Such nuclei can be formed either in bulk solution from HbS free monomers (the so-called homogeneous nucleation pathway), or on pre-existing polymers (the heterogeneous nucleation pathway). The model successfully describes the polymerization of fully deoxygenated HbS (i) in the 3-6 mM concentration (Ferrone et al 1985b), (ii) at different temperatures (15-35°C; Ferrone et al1985b), and (iii) in the presence of varying, clinically relevant, amounts of HbF (Rotter et al 2005a). …”

Section: The Double Nucleation Model Of Hbs Polymerizationmentioning

confidence: 99%

The Double Nucleation Model for Sickle Cell Haemoglobin Polymerization: Full Integration and Comparison with Experimental Data

Medkour¹,

Ferrone

Galactéros

et al. 2008

Acta Biotheor

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Sickle cell haemoglobin (HbS) polymerization reduces erythrocyte deformability, causing deleterous vaso-occlusions. The double-nucleation model states that polymers grow from HbS aggregates, the nuclei, (i) in solution (homogeneous nucleation), (ii) onto existing polymers (heterogeneous nucleation). When linearized at initial HbS concentration, this model predicts early polymerization and its characteristic delay-time (Ferrone et al. J Mol Biol 183(4):591-610, 611-631, 1985). Addressing its relevance for describing complete polymerization, we constructed the full, non-linearized model (Simulink), The MathWorks). Here, we compare the simulated outputs to experimental progress curves (n = 6-8 different [HbS], 3-6 mM range, from Ferrone's group). Within 10% from start, average root mean square (rms) deviation between simulated and experimental curves is 0.04 +/- 0.01 (25 degrees C, n = 8; mean +/- standard error). Conversely, for complete progress curves, averaged rms is 0.48 +/- 0.04. This figure is improved to 0.13 +/- 0.01 by adjusting heterogeneous pathway parameters (p < 0.01): the nucleus stability (sigma(2) micro( cc ): + 40%), and the fraction of polymer surface available for nucleation (phi), from 5e(-7), (3 mM) to 13 (6 mM). Similar results are obtained at 37 degrees C. We conclude that the physico-chemical description of heterogeneous nucleation warrants refinements in order to capture the whole HbS polymerization process.

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Molecular Crowding Limits the Role of Fetal Hemoglobin in Therapy for Sickle Cell Disease

Cited by 28 publications

References 32 publications

How can biochemical reactions within cells differ from those in test tubes?

How can biochemical reactions within cells differ from those in test tubes?

Inhibition of Hemoglobin S Polymerization in Vitro by a Novel 15-mer EF-Helix β73 Histidine-Containing Peptide

The Double Nucleation Model for Sickle Cell Haemoglobin Polymerization: Full Integration and Comparison with Experimental Data

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