In this section, we will develop novel technologies and approaches to increase the success rate of integral membrane protein structure determination by x-ray crystallography and cryo-electron microscopy (cryo-EM). The focus will be on sample preparation. Three of the four methods described depend on sequence optimization of the target membrane protein, either by (1) identifying and expanding upon regions of genomic sequence space most likely to harbor proteins amenable to in-depth structural analysis; (2) by producing libraries of random mutants of the target protein, and selecting those with increased probability of success in structure determination; (3) by genetically engineering chimeras between integral membrane proteins and soluble chaperones to promote either crystallization in lipidic cubic phase, or large molecular-weight assemblies for cryo-EM structure determination. In (4), we describe high-throughput screening methods to identify appropriate samples for high-resolution cryo-EM structure determination.
| Hartel, Andreas J W; Ong, Peijie; Schroeder, Indra et al. (2018) Single-channel recordings of RyR1 at microsecond resolution in CMOS-suspended membranes. Proc Natl Acad Sci U S A 115:E1789-E1798 |
| Blackburn, Matthew R; Hubbard, Caitlin; Kiessling, Volker et al. (2018) Distinct reaction mechanisms for hyaluronan biosynthesis in different kingdoms of life. Glycobiology 28:108-121 |
| Zhang, Yu; Kittredge, Alec; Ward, Nancy et al. (2018) ATP activates bestrophin ion channels through direct interaction. Nat Commun 9:3126 |
| Assur Sanghai, Zahra; Liu, Qun; Clarke, Oliver B et al. (2018) Structure-based analysis of CysZ-mediated cellular uptake of sulfate. Elife 7: |
| Su, Min; Gao, Feng; Yuan, Qi et al. (2017) Structural basis for conductance through TRIC cation channels. Nat Commun 8:15103 |
| Dufrisne, Meagan Belcher; Petrou, Vasileios I; Clarke, Oliver B et al. (2017) Structural basis for catalysis at the membrane-water interface. Biochim Biophys Acta Mol Cell Biol Lipids 1862:1368-1385 |
| Liu, Qun; Hendrickson, Wayne A (2017) Contemporary Use of Anomalous Diffraction in Biomolecular Structure Analysis. Methods Mol Biol 1607:377-399 |
| Scaglione, Antonella; Montemiglio, Linda Celeste; Parisi, Giacomo et al. (2017) Subcellular localization of the five members of the human steroid 5?-reductase family. Biochim Open 4:99-106 |
| Hirschi, Marscha; Johnson, Zachary Lee; Lee, Seok-Yong (2017) Visualizing multistep elevator-like transitions of a nucleoside transporter. Nature 545:66-70 |
| des Georges, Amédée; Clarke, Oliver B; Zalk, Ran et al. (2016) Structural Basis for Gating and Activation of RyR1. Cell 167:145-157.e17 |
Showing the most recent 10 out of 13 publications
