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G protein coupled receptors (GPCRs) mediate various of physiologicalactivities which makes them significant drug targets. Determination of atomic level structure of GPCRs facilitates the structure-based drug design. The most widely used method currently for solving GPCR structure is still protein crystallography especially lipidic

G protein coupled receptors (GPCRs) mediate various of physiologicalactivities which makes them significant drug targets. Determination of atomic level structure of GPCRs facilitates the structure-based drug design. The most widely used method currently for solving GPCR structure is still protein crystallography especially lipidic cubic phase (LCP) crystallization. LCP could mimic the native environment of membrane protein which stable the membrane proteins. Traditional synchrotron source requires large size large size protein crystals (>30 micron) due to the radiation damage during data collection. However, acquiring large sized protein crystals is challenging and not guaranteed practically. In this study, a novel method was developed which combined LCP technology and micro-electron diffraction (MicroED) technology. LCP-MicroED technology was able to collect complete diffraction data sets from serval submicron protein crystals and deliver high resolution protein structures. This technology was first confirmed with soluble protein crystals, proteinase K and small molecule crystals, cholesterol. Furthermore, this novel method was applied to a human GPCR target, Î22- adrenergic receptor (Î22AR). The structure model was successfully built which proved the feasibility of applying LCP-MicroED method to GPCRs and other membrane proteins. Besides, in this research, a novel human GPCR target, human histamine 4 receptor(H4R) was studied. Different constructs were expressed, purified, and characterized. Some key residuals that affect ligand binding were confirmed.
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    Title
    • Structure Determination of G Protein Coupled Receptor
    Contributors
    Date Created
    2022
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    Resource Type
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    • Partial requirement for: Ph.D., Arizona State University, 2022
    • Field of study: Biochemistry

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