多巴胺与人多巴胺转运蛋白在活细胞中相互作用的拉曼光谱特征标记,ACS Chemical Neuroscience

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多巴胺与人多巴胺转运蛋白在活细胞中相互作用的拉曼光谱特征标记
ACS Chemical Neuroscience ( IF 4.1 ) Pub Date : 2017-04-21 00:00:00 , DOI: 10.1021/acschemneuro.7b00048
Achut P. Silwal ₁ , Rajeev Yadav ₁ , Jon E. Sprague ₂ , H. Peter Lu ₁

Affiliation

多巴胺（DA）通过与人多巴胺转运蛋白（hDAT）和多巴胺受体的相互作用来控制中枢神经系统（CNS）的许多心理和行为活动。DA在中枢神经系统功能中的作用受药物与hDAT靶向结合的影响；因此，hDAT在神经生理学和神经病理生理学中起着至关重要的作用。一个有效的实验方法对于研究DA-hDAT的相互作用以及依赖于这种相互作用的多种药物（如精神刺激药和抗抑郁药）的作用是必要的。在寻找和识别拉曼光谱特征时，我们使用了表面增强拉曼散射（SERS）光谱来记录DA，人类胚胎肾293细胞（HEK293），hDAT-HEK293，DA-HEK293和DA-hDAT的SERS光谱-HEK293。我们已经展示了一种特定的2D分布SERS光谱分析方法来分析DA-hDAT相互作用。我们的研究表明，拉曼模式在807、839、1076、1090、1538和1665 cm^–1与DA–hDAT相互作用有关，其中拉曼位移在807和1076 cm处^–1是DA的结合状态的特征标记，用于探测DA–hDAT相互作用。根据密度函数理论（DFT）计算，在807 cm ^–1处DA束缚态的拉曼位移与弯曲模式α（C3–O10–H21），α（C2–O11–H22）的组合有关， α（C7–C8–H18），α（C6–C4–H13），α（C7–C8–H19）和α（C7–C8–N9）和拉曼位移在1076 cm ^–1处与弯曲模式α（H19–N9–C8），γ（N9–H19），γ（C8–H19），γ（N9–H20），γ（C8–H18）和α（C7–C8）的组合有关–H18）。这些发现表明，可以通过探测配体分子拉曼位移的变化来确认蛋白质-配体的相互作用，这对于理解神经递质及其受体或转运蛋白之间的分子相互作用可能至关重要。

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Raman Spectroscopic Signature Markers of Dopamine–Human Dopamine Transporter Interaction in Living Cells

Dopamine (DA) controls many psychological and behavioral activities in the central nervous system (CNS) through interactions with the human dopamine transporter (hDAT) and dopamine receptors. The roles of DA in the function of the CNS are affected by the targeted binding of drugs to hDAT; thus, hDAT plays a critical role in neurophysiology and neuropathophysiology. An effective experimental method is necessary to study the DA–hDAT interaction and effects of variety of drugs like psychostimulants and antidepressants that are dependent on this interaction. In searching for obtaining and identifying the Raman spectral signatures, we have used surface enhanced Raman scattering (SERS) spectroscopy to record SERS spectra from DA, human embryonic kidney 293 cells (HEK293), hDAT-HEK293, DA-HEK293, and DA-hDAT-HEK293. We have demonstrated a specific 2D-distribution SERS spectral analytical approach to analyze DA–hDAT interaction. Our study shows that the Raman modes at 807, 839, 1076, 1090, 1538, and 1665 cm^–1 are related to DA–hDAT interaction, where Raman shifts at 807 and 1076 cm^–1 are the signature markers for the bound state of DA to probe DA–hDAT interaction. On the basis of density function theory (DFT) calculation, Raman shift of the bound state of DA at 807 cm^–1 is related to combination of bending modes α(C3–O10–H21), α(C2–O11–H22), α(C7–C8–H18), α(C6–C4–H13), α(C7–C8–H19), and α(C7–C8–N9), and Raman shift at 1076 cm^–1 is related to combination of bending modes α(H19–N9–C8), γ(N9–H19), γ(C8–H19), γ(N9–H20), γ(C8–H18), and α(C7–C8–H18). These findings demonstrate that protein–ligand interactions can be confirmed by probing change in Raman shift of ligand molecules, which could be crucial to understanding molecular interactions between neurotransmitters and their receptors or transporters.

更新日期：2017-04-21

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