3st0
Engineered medium-affinity halide-binding protein derived from YFP: halide-freeEngineered medium-affinity halide-binding protein derived from YFP: halide-free
Structural highlights
FunctionGFP_AEQVI Energy-transfer acceptor. Its role is to transduce the blue chemiluminescence of the protein aequorin into green fluorescent light by energy transfer. Fluoresces in vivo upon receiving energy from the Ca(2+)-activated photoprotein aequorin. Publication Abstract from PubMedWe describe an engineered fluorescent optogenetic sensor, SuperClomeleon, that robustly detects inhibitory synaptic activity in single, cultured mouse neurons by reporting intracellular chloride changes produced by exogenous GABA or inhibitory synaptic activity. Using a cell-free protein engineering automation methodology that bypasses gene cloning, we iteratively constructed, produced, and assayed hundreds of mutations in binding-site residues to identify improvements in Clomeleon, a first-generation, suboptimal sensor. Structural analysis revealed that these improvements involve halide contacts and distant side chain rearrangements. The development of optogenetic sensors that respond to neural activity enables cellular tracking of neural activity using optical, rather than electrophysiological, signals. Construction of such sensors using in vitro protein engineering establishes a powerful approach for developing new probes for brain imaging. Visualization of synaptic inhibition with an optogenetic sensor developed by cell-free protein engineering automation.,Grimley JS, Li L, Wang W, Wen L, Beese LS, Hellinga HW, Augustine GJ J Neurosci. 2013 Oct 9;33(41):16297-309. doi: 10.1523/JNEUROSCI.4616-11.2013. PMID:24107961[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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