This grant application is focused on the question of how to incorporate the extensive basic knowledge of sickle hemoglobin polymerization into rational therapeutic approaches. 1. We will complete the description of the effect of mixtures on homogeneous and heterogenous nucleation of HbS, and test the description using laser photolysis methods. This will be critical for prediction of the outcome both of gene therapies and drug therapies, and will lay the groundwork for understanding mixtures of liganded and unliganded HbS. 2. As an approach to therapy, we will extend the double nucleation model to include liquid-liquid phase- transitions and their effect on polymer formation. Because of the dramatic effects of crowding (which we have previously measured), the formation of dense liquid regions lowers the Hb concentration elsewhere. This approach offers as much as a tenfold improvement over the therapeutic potential of inducing Fetal hemoglobin, which is the foundation of present hydroxyureatherapy. The effect has already been seen in our preliminary work. Photolysis tests will also determine if nucleation and phase separation are correctly combined. 3. We will continue measurements of fiber flexibility and domain rigidity begun in the last grant period, (a) We are measuring the rigidity of polymer domains by magnetically driven beads and simultaneous optical imagining of the growing domains (b) We are measuring fiber rigidty using patterned photolysis to create bent optical channels through which fibers can grow. A number of protein association diseases are known, though none are so well characterized as that of HbS. As a result, the types of questions that can be raised about the biophysical mechanism have often been raised first for HbS, and their solutions then become available to study other diseases as well. Hence, successes generated here are expected to have useful spin-offs to other protein association problems.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Macromolecular Structure and Function B Study Section (MSFB)
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Luksenburg, Harvey
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Drexel University
Schools of Arts and Sciences
United States
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Yosmanovich, Donna; Rotter, Maria; Aprelev, Alexey et al. (2016) Calibrating Sickle Cell Disease. J Mol Biol 428:1506-14
Tokarev, Alexander; Aprelev, Alexey; Zakharov, Mikhail N et al. (2012) Multifunctional magnetic rotator for micro and nanorheological studies. Rev Sci Instrum 83:065110
Weng, Weijun; Ferrone, Frank A (2011) Metastable gels: A novel application of Ogston theory to sickle hemoglobin polymers. Biophys Chem 154:99-101
Rotter, Maria; Yosmanovich, Donna; Briehl, Robin W et al. (2011) Nucleation of sickle hemoglobin mixed with hemoglobin A: experimental and theoretical studies of hybrid-forming mixtures. Biophys J 101:2790-7
Rotter, Maria A; Chu, Haiyan; Low, Philip S et al. (2010) Band 3 catalyzes sickle hemoglobin polymerization. Biophys Chem 146:55-9
Zakharov, Mikhail N; Aprelev, Alexey; Turner, Matthew S et al. (2010) The microrheology of sickle hemoglobin gels. Biophys J 99:1149-56
Weng, Weijun; Aprelev, Alexey; Briehl, Robin W et al. (2008) Universal metastability of sickle hemoglobin polymerization. J Mol Biol 377:1228-35
Aprelev, Alexey; Weng, Weijun; Zakharov, Mikhail et al. (2007) Metastable polymerization of sickle hemoglobin in droplets. J Mol Biol 369:1170-4
Adachi, Kazuhiko; Ding, Min; Surrey, Saul et al. (2006) The Hb A variant (beta73 Asp-->Leu) disrupts Hb S polymerization by a novel mechanism. J Mol Biol 362:528-38
Rotter, Maria A; Aprelev, Alexey; Dragos, Dana et al. (2004) Aspartame has no effect on the polymerization of sickle hemoglobin. Clin Pharmacol Ther 75:248-9

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