Modeling of the structure and functional mechanisms of voltage-gated channels

电压门控通道的结构和功能机制建模

• 批准号: 7592879
• 负责人: HOMER ROBERT GUY
• 金额: $ 27.86万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7592879
• 关键词: Binding; Calculi; Categories; Complex; Computing Methodologies; Cyclic Nucleotides; Cysteine; Cytoplasmic Tail; Data; Databases; Drug Design; Electron Spin Resonance Spectroscopy; Evolution; Family; Gene Targeting; Geum; Glutamates; Goals; Hereditary Disease; Homology Modeling; Human; Ions; Kv1.2' channel; Lipid Bilayers; Malignant Neoplasms; Manuscripts; Membrane Proteins; Methods; Modeling; Molecular; Molecular Conformation; Movement; Mutagenesis; Mutation; Paramecium; Pharmacologic Substance; Pharmacology; Plants; Potassium Channel; Property; Proteins; Proto-Oncogene Proteins c-akt; Rest; Roentgen Rays; Scanning; Scorpions; Simulate; Structural Models; Structure; Testing; Thermodynamics; Toxin; Transmembrane Domain; channel blockers; computer studies; extracellular; hypertensive heart disease; molecular dynamics; sensor; symporter; voltage; voltage gated channel

During the last two years, we have developed structural models of the transmembrane and extracellular segments of Shaker, KvAP, hERG, NaChBac, and Ca2+ channels in resting, open, and numerous transition conformations. Molecular dynamic simulations of these channels embedded in a lipid bilayer were performed to evaluate and refine the models. The models were constrained by recently obtained experimental data; e.g., the crystal structure of the Kv1.2 channel, electron paramagnetic resonance (EPR) studies of KvAP channels, thermodynamic cyclic mutagenesis studies of the binding of BeKM1 toxin from scorpions to the hERG channel, and cysteine scanning mutagenesis (SCAM) studies of Ca2+ channel pores. We have demonstrated that the helical screw model for the voltage-dependent movement of the S4 voltage-sensor segment that we proposed first in 1986, is consistent with virtually all experimental results and energetic criteria, including analyses using molecular dynamic simulations. Recent experimental and computational studies from other groups have provided additional support for our models. The NaChBac channel is a prokaryotic Na+ channel that has similarities to K+, Ca2+, and Na+ channels. We were the first group to identify this sequence in the prokaryotic sequence data base. Since then, it has been expressed and its properties have been studied expensively. Efforts are underway to solve its crystal structure. Our NaChBac was develop using the crystal structure of the Kv1.2 channel as an initial template. The resulting NaChBac model has several unique features involving the ion selective region formed by the P segments, the activation gate formed by the S6 segment, and the interaction between the voltage-sensing (S1-S4) and pore-forming (S5-P-S6) domains. We are now using the NaChBac model as a stepping stone to model more complex eukaryotic Ca2+ and Na+ channels. So far we have modeled the transmembrane regions of human and fungal Ca2+ channels. We are collaborating with Steffen Herrings and Angie Gellis groups to test experimentally these models. Speciffically, we are using the models to analyze the molecular pharmacology of Ca2+ channel blockers (important in treating hypertension and heart disease in humans and potentially important as antifungicides) and to better understand how mutations associated with genetic diseases alter the gating properties of Ca2+ channels. Manuscripts have been submitted on our models of NaChBac and human Ca2+ channels. We have started a new project to develop structural and functional models of channel families [EAG and ERG, AKT (plant), PAK (paramecium), CNG, and HCN] that possess a cyclic nucleotide domain. Substantial progress has been made in modeling the hERG and HCN channels. A crystal structure of the cyclic nucleotide-binding domain of the HCN channel is being used to model the cytoplasmic domain. We are focusing on this group of families because expression of some of them (especially EAG channels that are closely related to hERG channels) has been associated with several cancers. We are collaborating with Gea-Ny Tsengs lab to experimentally test aspects of our hERG channels.

在过去的两年中，我们已经开发了跨膜的结构模型， Shaker，KvAP，hERG，NaChBac和Ca 2+通道的细胞外片段在静息，开放和 许多过渡构象。这些通道的分子动力学模拟嵌入在 进行脂质双层以评估和改进模型。模型受到约束 通过最近获得的实验数据;例如，Kv1.2通道的晶体结构， KvAP通道的电子顺磁共振（EPR）研究，热力学循环 来自蝎子的BeKM 1毒素与hERG通道结合的诱变研究，以及 半胱氨酸扫描诱变（SCAM）研究Ca 2+通道孔。我们已经证明 S4的电压依赖运动的螺旋模型 我们在1986年首次提出的电压传感器部分，几乎与所有 实验结果和能量标准，包括分子动力学分析 模拟最近来自其他小组的实验和计算研究提供了 为我们的模型提供更多支持。NaChBac通道是一种原核Na+通道， 与K+、Ca 2+和Na+通道相似。我们是第一个发现这个序列的小组 在原核生物序列数据库中。从那时起，它就被表达出来， 已经被昂贵地研究过了。目前正在努力解决其晶体结构。我们 使用Kv1.2通道的晶体结构作为初始模板开发NaChBac。 由此产生的NaChBac模型具有涉及离子选择性区域的几个独特特征 由P段形成的激活栅极、由S6段形成的激活栅极以及由P段形成的相互作用栅极。 在电压敏感域（S1-S4）和成孔域（S5-P-S6）之间。我们现在使用 NaChBac模型是模拟更复杂的真核Ca 2+和Na+通道的垫脚石。所以 到目前为止，我们已经模拟了人类和真菌Ca 2+通道的跨膜区域。我们 与Steffen Herrings和Angie Gellis团队合作， 这些模型的实验。具体地说，我们正在使用模型来分析分子 Ca 2+通道阻滞剂的药理学（在治疗高血压和心脏病方面很重要， 人类和潜在的重要的抗真菌剂），并更好地了解突变如何 与遗传性疾病相关的钙通道改变了钙通道的门控特性。手稿 已经提交了关于我们的NaChBac和人类Ca 2+通道模型的研究报告。我们开始了一个新的 开发通道家族的结构和功能模型的项目[EAG和ERG，AKT （植物）、PAK（草履虫）、CNG和HCN]。实质性 在对hERG和HCN通道建模方面已经取得了进展。的晶体结构 HCN通道的环核苷酸结合结构域被用于模拟细胞质 域我们之所以关注这类家庭，是因为他们中的一些人 （特别是与hERG通道密切相关的EAG通道）与 几种癌症。我们正在与Gea-Ny Tsengs实验室合作， 我们的hERG通道。