The goal is to understand the molecular mechanisms of sugar transport by transmembrane proteins. Transport is largely achieved by a family of proteins which span the membrane 12 times. Two transporters have been selected from this family: the human erythrocyte glucose transporter (Glut1) and the E. coli glucuronide transporter (GusB). Glut 1 is a ubiquitous facilitative glucose transport protein in humans, abundant in erythrocytes and in the brain, providing these tissues with glucose, their primary energy source. GusB is the protein responsible for transporting glucuronide sugars into the E. Coli bacterium in the human intestine.
The aim i s to determine the structure of these proteins in order to understand their transport mechanisms. Large, highly ordered two- or three-dimensional crystals are required for structural determination of transmembrane proteins. Such crystals can only be grown from protein-lipid-detergent complexes that are homogenous. To this end, the investigators have purified Glut1 and GusB in ten-milligram quantities, and have found conditions to keep these proteins active, stable, and monodisperse. They are applying and developing novel methods for their crystallization. Cryo-electron microscopy, electron diffraction, and x-ray crystallography will be employed to determine the protein structure from such crystals. Knowledge of the structure of these proteins and their conformational changes will reveal their transport mechanisms. In addition, these structures will become paradigms for other members of the 12-transmembrane helix transporter family, including the glucose transporter in muscle and fat tissues (Glut4), which is involved in the pathogenesis of type II diabetes, and the dopamine transporter responsible for cocaine addiction.
| Karpowich, Nathan K; Song, Jinmei; Wang, Da-Neng (2016) An Aromatic Cap Seals the Substrate Binding Site in an ECF-Type S Subunit for Riboflavin. J Mol Biol 428:3118-30 |
| Sauer, David B; Karpowich, Nathan K; Song, Jin Mei et al. (2015) Rapid Bioinformatic Identification of Thermostabilizing Mutations. Biophys J 109:1420-8 |
| Karpowich, Nathan K; Song, Jin Mei; Cocco, Nicolette et al. (2015) ATP binding drives substrate capture in an ECF transporter by a release-and-catch mechanism. Nat Struct Mol Biol 22:565-71 |
| Mulligan, Christopher; Fitzgerald, Gabriel A; Wang, Da-Neng et al. (2014) Functional characterization of a Na+-dependent dicarboxylate transporter from Vibrio cholerae. J Gen Physiol 143:745-59 |
| Loew, Leslie M; Wang, Da-Neng (2013) Science communication: Quality at stake. Science 342:1169 |
| Wang, Da-Neng; Stieglitz, Heather; Marden, Jennifer et al. (2013) Benjamin Franklin, Philadelphia's favorite son, was a membrane biophysicist. Biophys J 104:287-91 |
| Waight, Andrew B; Czyzewski, Bryan K; Wang, Da-Neng (2013) Ion selectivity and gating mechanisms of FNT channels. Curr Opin Struct Biol 23:499-506 |
| Karpowich, Nathan K; Wang, Da-Neng (2013) Assembly and mechanism of a group II ECF transporter. Proc Natl Acad Sci U S A 110:2534-9 |
| Mancusso, Romina; Gregorio, G Glenn; Liu, Qun et al. (2012) Structure and mechanism of a bacterial sodium-dependent dicarboxylate transporter. Nature 491:622-6 |
| Czyzewski, Bryan K; Wang, Da-Neng (2012) Identification and characterization of a bacterial hydrosulphide ion channel. Nature 483:494-7 |
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