This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. This project involves three different and unique integral membrane proteins: (1) a bacterial transport protein ('X4');(2) a eukaryotic P2X ion channel;and (3) a eukaryotic, full-length ionotropic glutamate receptor (iGluR). For all three projects, the scientific or technical purpose is the same ?to measure the highest resolution diffraction data sets possible. This presents a great challenge because all these crystals diffract weakly ?they are crystals of integral membrane proteins and a large fraction of the crystal volume is composed of aqueous solution and disordered detergent and lipid molecules. In particular, for the X4 project, we are aiming to collect high resolution native data on the NE-CAT microfocus beam line and multiwavelength data on the ID-24 beam line. For the P2X project, since we have low resolution phases, we are measuring native data sets (with and without agonists and modulators) on the microfocus beam line. In the case of the glutamate receptor project, we are using the known structures of the extracellular domains in molecular replacement to obtain phase information, and thus the emphasis is on measuring single wavelength data on a large number of agonist, antagonist and allosteric modulator complexes. The importance of all three projects is substantial. To the best of my knowledge, the folds for the X4 and P2X proteins are currently unknown, i.e. the determined folds are likely to be unique, and thus these initial structures will provide the architectural blueprint for two large and very important classes of membrane proteins. The detailed structural analysis, in addition, will provide deep and as-of-yet unknown insights into molecular mechanism. In the case of the glutamate receptor, the structure of the intact receptor will provide profound insights into the mechanism of ion channel gating and its modulation, as well as into ion permeation and block.
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