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Recombinant expression and purification of pentameric ligand-gated ion channels for Cryo-EM structural studies

Methods Enzymol. 2021-02; 
Arvind Kumar, Sandip Basak, Sudha Chakrapani
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Gene Synthesis … 3) were codon optimized forSf9cells (Spodoptera frugiperda) and synthesized at GenScript USA Inc?… inserted at the C-terminus of 5-HT3AR gene (Fig?… Finally, the genes were insertedintopFastBac1vector (Invitrogen) whichisthenusedforrecombinant baculovirus generation?… Get A Quote

摘要

Pentameric ligand-gated ion channels (pLGICs) are central players in synaptic neurotransmission and are targets to a range of drugs used to treat neurological disorders and pain. pLGICs are intrinsically dynamic membrane proteins that upon stimulation by neurotransmitters, undergo global conformational changes across multiple domains spanning a distance of over 165?. The inter-domain flexibility, a feature crucial for their function as signal transducers in chemical synapses, has been problematic in the efforts toward determining high-resolution structures. Earlier structural studies tackled this issue with a variety of strategies that included partial truncation of flexible domains and the use of antibodies a... More

關鍵詞

Cryo-EM, Cys-loop receptors, Gating, Nanodisc, Reconstitution, pLGIC
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