Targeting cell surface receptors with ligand-conjugated nanocrystals
Targeting cell surface receptors with ligand-conjugated nanocrystals
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DOI：
10.1021/ja003486s
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发表时间：
2002-05-01
影响因子：
15
通讯作者：
Blakely, RD
中科院分区：
文献类型：
--
作者：
Rosenthal, SJ;Tomlinson, A;Blakely, RD
为了探索使用配体缀合的纳米晶体靶向细胞表面受体、离子通道和转运蛋白的潜力，我们探索了阿托洛宁标记的CdSe纳米晶体（SNAC）与抗抑郁药敏感的人和果蝇5-羟色胺转运蛋白（hSERT，dSERT）在HeLa和HEK-293细胞中表达的相互作用的能力。与未缀合的纳米晶体不同，发现SNAC剂量依赖性地抑制hSERT和dSERT对放射性标记的5-羟色胺的转运，估计半最大活性（EC 50）为33（dSERT）和99 μ M（hSERT）。当5-羟色胺通过连接臂（LSNAC）缀合到hSERT时，测定hSERT的EC 50为115 μ M。电生理学测量表明，LSNAC不会从5-羟色胺-3（5-HT（3））受体引发电流，但当暴露于转运蛋白时确实会产生电流，这与拮抗剂引发的电流相似。此外，发现荧光LSNAC标记SERT转染的细胞，但不标记未转染的细胞或与高亲和力SERT拮抗剂帕罗西汀共孵育的转染细胞。这些发现支持进一步考虑配体共轭纳米晶体作为活细胞中膜蛋白的通用探针。
To explore the potential for use of ligand-conjugated nanocrystals to target cell surface receptors, ion channels, and transporters, we explored the ability of serotonin-labeled CdSe nanocrystals (SNACs) to interact with antidepressant-sensitive, human and Drosophila serotonin transporters (hSERT, dSERT) expressed in HeLa and HEK-293 cells. Unlike unconjugated nanocrystals, SNACs were found to dose-dependently inhibit transport of radiolabeled serotonin by hSERT and dSERT, with an estimated half-maximal activity (EC50) of 33 (dSERT) and 99 muM (hSERT). When serotonin was conjugated to the nanocrystal through a linker arm (LSNACs), the EC50 for hSERT was determined to be 115 muM. Electrophysiology measurements indicated that LSNACs did not elicit currents from the serotonin-3 (5HT(3)) receptor but did produce currents when exposed to the transporter, which are similar to those elicited by antagonists. Moreover, fluorescent LSNACs were found to label SERT-transfected cells but did not label either nontransfected cells or transfected cells coincubated with the high-affinity SERT antagonist paroxetine. These findings support further consideration of ligand-conjugated nanocrystals as versatile probes of membrane proteins in living cells.