Project is concerned with structure determination by 2-D crystallization on lipid monolayers of Myelin Basic Protein (MBP), an 18.5 kDa protein that is a major constituent of nerve myelin. Myelin is a multi-layered structure surrounding a nerve that not only provides electrical insulation, but by virtue of its highly compact structure and resultant low capacitance, enables the fast propagation of nerve impulses necessary to the functioning of higher organisms. MBP is believed to be essential to the compaction of nerve myelin. As the primary antigenic determinant in Multiple Sclerosis (MS), the critical role of MBP is illustrated by the devastating neurological consequences of the disease. Using the fact that his protein is known to bind strongly to acidic phospholipids, we have employed the """"""""lipid layer"""""""" method of Uzgiris and Kornberg (1983) to get a variety of 2-D assemblies. We have found that the formation of crystals depends strongly on the head group of the surfactant used. Our best results have been obtained with a derivatized 10, 12-Pentacosadiynoic acid and we have obtained crystals that diffract to better than 30 Angstroms. A low resolution 3-D structure of MBP determined by single particle methods has been recently published (Benaic et al., 1997). While they describe a 110 Angstroms diameter particles with a shape roughly in agreement with the approximately 80 Angstroms repeating unit seen in our unit cell, they attribute this particle to a single molecule of MBP. We believe, from basic physical considerations, that his in incorrect and that the particle is a multimer of at least 6 molecules. We will perform a tilt series on crystals of MBP to create a 3-D reconstruction that will resolve this issue. Following this, we will extend the resolution by improving the purification of this protein.
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| Nogales, Eva (2018) Cryo-EM. Curr Biol 28:R1127-R1128 |
| Sazzed, Salim; Song, Junha; Kovacs, Julio A et al. (2018) Tracing Actin Filament Bundles in Three-Dimensional Electron Tomography Density Maps of Hair Cell Stereocilia. Molecules 23: |
| Kamennaya, Nina A; Zemla, Marcin; Mahoney, Laura et al. (2018) High pCO2-induced exopolysaccharide-rich ballasted aggregates of planktonic cyanobacteria could explain Paleoproterozoic carbon burial. Nat Commun 9:2116 |
| Howes, Stuart C; Geyer, Elisabeth A; LaFrance, Benjamin et al. (2018) Structural and functional differences between porcine brain and budding yeast microtubules. Cell Cycle 17:278-287 |
| Glaeser, Robert M (2018) PROTEINS, INTERFACES, AND CRYO-EM GRIDS. Curr Opin Colloid Interface Sci 34:1-8 |
| Kellogg, Elizabeth H; Hejab, Nisreen M A; Poepsel, Simon et al. (2018) Near-atomic model of microtubule-tau interactions. Science 360:1242-1246 |
| Zhang, Rui; LaFrance, Benjamin; Nogales, Eva (2018) Separating the effects of nucleotide and EB binding on microtubule structure. Proc Natl Acad Sci U S A 115:E6191-E6200 |
| Nogales, Eva (2018) Cytoskeleton in high resolution. Nat Rev Mol Cell Biol 19:142 |
| Han, Bong-Gyoon; Watson, Zoe; Cate, Jamie H D et al. (2017) Monolayer-crystal streptavidin support films provide an internal standard of cryo-EM image quality. J Struct Biol 200:307-313 |
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