Distinct Structural Features of Cyclothiazide Are Responsible for Effects on Peak Current Amplitude and Desensitization Kinetics at iGluR2
Distinct Structural Features of Cyclothiazide Are Responsible for Effects on Peak Current Amplitude and Desensitization Kinetics at iGluR2
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DOI：
10.1016/j.jmb.2009.07.002
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发表时间：
2009-09-04
影响因子：
5.6
通讯作者：
Kastrup, Jette S.
中科院分区：
文献类型：
--
作者：
Hald, Helle;Ahring, Philip K.;Kastrup, Jette S.
离子型谷氨酸受体(IGluRs)介导快速兴奋性神经传递。当应用谷氨酸时，2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic酸受体经历快速且几乎完全的脱敏，这种脱敏可以被正变构调节剂减弱。正变构调节的分子机制已经通过iGluR2的配体结合核心的晶体结构来阐明，例如与环噻嗪(CTZ)的络合物。在这里，我们用X射线结晶学和快速应用膜片钳电生理的方法研究了CTZ和三个与其密切相关的类似物NS1493、NS5206和NS5217在iGluR2上的结构和功能。CTZ是四种化合物中对iGluR2(Q)(I)[对谷氨酸反应的E-max：754%(CTZ)，490%(NS1493)，399%(NS5206)和476%(NS5217)，EC50：10(CTZ)，26(NS1493)，43(NS5206)和48(NS5217)]中最有效和最有效的调节剂。根据药效和脱敏动力学，四种调节剂分为三组：(1)CTZ使峰值电流效应增加一倍，几乎完全阻断受体脱敏；(2)NS5206和NS5217低效，仅部分减弱脱敏；(3)NS1493低效，但几乎完全阻断受体脱敏。1，1-dioxo-3，4-dihydro-2H-benzo[e][1，2，4]thiadiazine环系3-位上的疏水取代基对化合物的药效很重要，顺序如下：降冰片烯基(bicyclo[2.2.1]hept-2-ene)>；cyclopentyl>；methyl.用疏水性显著降低的环戊烷环取代降冰片烯基增加了调节剂的灵活性，因为环戊烷环在iGluR2变构结合部位采用了不同的构象。导致几乎完全脱敏的主要结构特征是在1，1-dioxo-3，4-dihydro-2H-benzo[e][1，2，4]thiadiazine环系统的4-位存在一个NH氢键供体，形成一个锚定在Ser754上的氢键。因此，CTZ 4位上的原子似乎是受体脱敏动力学的主要决定因素。(C)2009爱思唯尔有限公司。保留所有权利。
Ionotropic glutamate receptors (iGluRs) mediate fast excitatory neurotransmission. Upon glutamate application, 2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid receptors undergo rapid and almost complete desensitization that can be attenuated by positive allosteric modulators. The molecular mechanism of positive allosteric modulation has been elucidated previously by crystal structures of the ligand-binding core of iGluR2 in complex with, for example, cyclothiazide (CTZ). Here, we investigate the structure and function of CTZ and three closely related analogues NS1493, NS5206, and NS5217 at iGluR2, by X-ray crystallography and fast application patch-clamp electrophysiology. CTZ was the most efficacious and potent modulator of the four compounds on iGluR2(Q)(i) [E-max normalized to response of glutamate: 754% (CTZ), 490% (NS1493), 399% (NS5206), and 476% (NS5217) and EC50 in micromolar: 10 (CTZ), 26 (NS1493),43 (NS5206), and 48 (NS5217)]. The four modulators divide into three groups according to efficacy and desensitization kinetics: (1) CTZ increases the peak current efficacy twice as much as the three analogues and nearly completely blocks receptor desensitization; (2) NS5206 and NS5217 have low efficacy and only attenuate desensitization partially; (3) NS1493 has low efficacy but nearly completely blocks receptor desensitization. A hydrophobic substituent at the 3-position of the 1,1-dioxo-3,4-dihydro-2H-benzo[e][1,2,4]thiadiazine ring system is important for compound efficacy, with the following ranking: norbornenyl (bicyclo[2.2.1]hept-2-ene)>cyclopentyl>methyl. The replacement of the norbornenyl moiety with a significantly less hydrophobic cyclopentane ring increases the flexibility of the modulator as the cyclopentane ring adopts various conformations at the iGluR2 allosteric binding site. The main structural feature responsible for a nearly complete block of desensitization is the presence of an NH hydrogen bond donor in the 4-position of the 1,1-dioxo-3,4-dihydro-2H-benzo[e][1,2,4]thiadiazine ring system, forming an anchoring hydrogen bond to Ser754. Therefore, the atom at the 4-position of CTZ seems to be a major determinant of receptor desensitization kinetics. (C) 2009 Elsevier Ltd. All rights reserved.