| 题名 | 电压门控钾离子通道天然小分子调节剂的筛选与机制研究 |
| 作者 | 叶臣君 |
| 学位类别 | 硕士 |
| 答辩日期 | 2015-05 |
| 授予单位 | 中国科学院研究生院 |
| 授予地点 | 北京 |
| 导师 | 周鸣 ,杨建 |
| 关键词 | 电压门控钾离子通道 Kv1 天然小分子 TEVC 内面向外式膜片钳 |
| 其他题名 | Screening and study of natural molecule modulators of voltage-gated potassium channels |
| 中文摘要 | 电压门控钾离子通道在神经系统、心脏和免疫系统中发挥着重要作用。中药在治疗心血管、神经系统及免疫相关疾病方面有确切的疗效，但是中药成分复杂，有效分子、作用靶点及如何作用等治疗疾病的具体机理不清楚。种类繁多的中药就是一个蕴藏着许多对离子通道有调节作用分子的宝藏，等待我们去开采。 本论文以中药和中药来源的天然小分子为筛选资源，以Kv1通道为筛选靶点，采用TEVC双电极电压钳技术筛选对Kv1有作用的激活剂或者抑制剂。中药粗提物筛选结果表明，75种中药中有3种对Kv1通道有明显的抑制作用；小分子筛选结果表明，180种单体分子中成功筛选出8种对Kv1通道有强烈作用的分子。 我们选择来自南欧大戟的Kwls2和Kwls10以及来自灵芝的Kfb18分子，采用TEVC双电极电压钳技术和内面向外式膜片钳技术进一步研究。结果如下： （1）Kwls2对Kv1家族的抑制作用有相对于Kv3家族和T型钙通道的特异性； Kwls2浓度依赖性地抑制Kv1.1通道，其IC50为4.78±1.17 μM。 （2）Kwls10对Kv1.3、Kv1.5和Kv1.1亚型的抑制作用有相对于Kv1.4亚型的特异性； Kwls10浓度依赖性地抑制Kv1.3通道，其IC50为9.5±1.0 μM； Kwls10很可能是结合于Kv1.3通道的细胞膜内侧。 （3）Kfb18对Kv1.3和Kv1.4有特异抑制作用，而对Kv1.1、Kv1.5、Kv2.1和Kv3.1都没有抑制作用；Kfb18分子可能结合于Kv1.3通道的细胞膜外侧而发挥抑制作用。 （4）Kfb18对Kv1.1的激活作用表现为，使Kv1.1通道激活曲线左移，更容易打开，并且左移作用具有浓度依赖性。 （5）测试一系列Kfb18类似物对Kv1的作用，并比对Kfb18及类似物的结构，得到了Kfb18表现双重作用的重要分子信息。 本论文发现了对Kv1通道有调节作用的多种天然小分子，为解释中药药理作用提供了线索，也为开发新型Kv1通道调节剂提供了宝贵的分子模板。 |
| 英文摘要 | Voltage-gated potassium channels play a vital role in nervous system, heart and immune sysytem. Herbs have clear therapeutic effect on related diseases, but herbs have so complicated components that little is known about their effective molecules, their targets and how they function in detail, thus herbs of a great variety is like an unearth buried treasure for ion channel modulators, waiting for us to dig deeper. With herb extracts and small molecules from herbs as resources for screening, and Kv1 channels as screening targets, we aimed to discover new Kv1 channel openers or inhibitors by method of TEVC( Two-electrode voltage clamp). Screening results are as follows, there are 3 out of 75 herb extracts and 8 out of 180 small moleculess showing great effect on Kv1 channel. We choosed Kwls2 and Kwls10 from Euphorbia peplus and Kfb18 from ganoderma for further study with the method of TEVC( Two-electrode voltage clamp) and inside-out patch. Results are as follows: (1)Kwls2 inhibits Kv1 channels selectively over Kv3 channels and T type calcium channel, and the inhibition is concentration dependent, with an IC50 of 4.78±1.17 μM for Kv1.1. (2)Kwls10 inhibits Kv1.1, Kv1.3 and Kv1.5 selectively over Kv1.4, and the inhibition is concentration dependent, with an IC50 of 9.5±1.0 μM for Kv1.3. Kwls10 probably binds the cytoplasma side of Kv1.3 channel. (3)Kfb18 inhibits Kv1.3 and Kv1.4 selectively over Kv1.1, Kv1.5, Kv2.1 and Kv3.1 and Kfb18 probably binds the extracellular side of Kv1.3 channel. (4)Kfb18 activates Kv1.1 channel by left shifting the activation curve, making it easier for Kv1.1 channel to open, and this effect is also concentration dependent. (5)We got some important informations about which parts are important for Kfb18's dual function by comparing the strucure and effect of Kfb18 analogues. In conclusion, we find several natural small molecules as Kv1 channel modulators, which providing clues for explanation of herbs' pharmacological activities and offering valuable molecular templets for developing of novel Kv1 channel modulators. |
| 语种 | 中文 |
| 内容类型 | 学位论文 |
| 源URL | [http://159.226.149.26:8080/handle/152453/10125]  |
| 专题 | 昆明动物研究所_离子通道药物研发中心 |
| 推荐引用方式 GB/T 7714 | 叶臣君. 电压门控钾离子通道天然小分子调节剂的筛选与机制研究[D]. 北京. 中国科学院研究生院. 2015. |
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