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J Am Chem Soc
2006 Dec 13;12849:15701-13. doi: 10.1021/ja064324k.
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Binding of muscimol-conjugated quantum dots to GABAC receptors.
Gussin HA
,
Tomlinson ID
,
Little DM
,
Warnement MR
,
Qian H
,
Rosenthal SJ
,
Pepperberg DR
.
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Functionalization of highly fluorescent CdSe/ZnS core-shell nanocrystals (quantum dots, qdots) is an emerging technology for labeling cell surface proteins. We have synthesized a conjugate consisting of approximately 150-200 muscimols (a GABA receptor agonist) covalently joined to the qdot via a poly(ethylene glycol) (PEG) linker (approximately 78 ethylene glycol units) and investigated the binding of this muscimol-PEG-qdot conjugate to homomeric rho1 GABAC receptors expressed in Xenopus oocytes. GABAC receptors mediate inhibitory synaptic signaling at multiple locations in the central nervous system (CNS). Binding of the conjugate was analyzed quantitatively by determining the fluorescence intensity of the oocyte surface membrane in relation to that of the surrounding incubation medium. Upon 5- to 10-min incubation with muscimol-PEG-qdots (34 nM in qdot concentration), GABAC-expressing oocytes exhibited a fluorescent halo at the surface membrane that significantly exceeded the fluorescence of the incubation medium. This halo was absent following muscimol-PEG-qdot treatment of oocytes lacking GABAC receptors. Incubation of the oocyte with free muscimol (100 microM-5 mM), PEG-muscimol (500 microM), or GABA (100 microM - 5 mM) substantially reduced or eliminated the fluorescence halo produced by muscimol-PEG-qdots, and the removal of GABA or free muscimol led to a recovery of muscimol-PEG-qdot binding. Unconjugated qdots and PEG-qdots that lacked conjugated muscimol neither exhibited significant binding activity nor diminished the subsequent binding of muscimol-PEG-qdots. The results indicate that muscimol joined to qdots via a long-chain PEG linker exhibits specific binding activity at the ligand-binding pocket of expressed GABAC receptors, despite the presence of both the long PEG linker and the sterically bulky qdot.
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17147380
???displayArticle.pmcLink???PMC2553244 ???displayArticle.link???J Am Chem Soc ???displayArticle.grants???[+]
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