Lipid Bilayers
Lipid Bilayers
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脂质双层
DOI：
--
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
Dr. Thomas Gutberlet
中科院分区：
文献类型：
--
作者：
Dr. John Katsaras;Dr. Thomas Gutberlet
大电导钙激活钾（Kca）通道从天然细胞膜到平面脂质双层的重建提供了一个强大的方法来研究单通道特性，包括离子传导，药理学和门控。最近，来自果蝇Slowpoke（Slo）基因的Kca通道已被克隆并在非洲爪蟾卵母细胞中异源表达。在这份报告中，我们描述了重建的克隆和表达的SLO KCA通道从非洲爪蟾卵母细胞膜成脂质双层。重建的通道显示出天然KCa通道的功能特性。它们在对称钾（250 mM）中具有-260 pS的平均单位电导，并且它们是电压和钙敏感的。在50 pM Ca 2+时，它们的半活化电位接近-20 mV;它们对钙的亲和力在微摩尔范围内。重构的Slo Kca通道对外部Charybdotoxin（40-500 nM）不敏感，对外部四乙基铵（KD = 158 pM，0 mV）和内部Ba 2+（KD = 76 pM，40 mV）的微摩尔浓度敏感。此外，它们被内部应用的“球”灭活肽（KD = 480 pM，40 mV）阻断。这些结果表明，克隆的KCA通道表达在非洲爪蟾卵母细胞可以很容易地纳入脂双层，其中详细的机制研究，可以在受控的内部和外部的实验条件下进行。
Reconstitution of large conductance calcium-activated potassium (Kca) channels from native cell membranes into planar lipid bilayers provides a powerful method to study single channel properties, including ion conduction, pharmacology, and gating. Recently, Kca channels derived from the Drosophila Slowpoke (Slo) gene have been cloned and heterologously expressed in Xenopus oocytes. In this report, we describe the reconstitution of cloned and expressed Slo Kca channels from Xenopus oocyte membranes into lipid bilayers. The reconstituted channels demonstrate functional properties characteristic of native Kca channels. They possess a mean unitary conductance of -260 pS in symmetrical potassium (250 mM), and they are voltageand calcium-sensitive. At 50 pM Ca2+, their half-activation potential was near -20 mV; and their affinity for calcium is in the micromolar range. Reconstituted Slo Kca channels were insensitive to external charybdotoxin (40-500 nM) and sensitive to micromolar concentrations of external tetraethylammonium (KD = 158 pM, at 0 mV) and internal Ba2+ (KD = 76 pM, at 40 mV). In addition, they were blocked by internally applied "ball" inactivating peptide (KD = 480 pM, at 40 mV). These results demonstrate that cloned Kca channels expressed in Xenopus oocytes can be readily incorporated into lipid bilayers where detailed mechanistic studies can be performed under controlled internal and external experimental conditions.
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膜蛋白的核磁共振结构研究。
影响因子：
6.8
作者：
Marassi,FM;Opella,SJ
通讯作者：
Opella,SJ
定向膜蛋白的 NMR 数据模拟：实验设计的实用信息。
影响因子：
3.4
作者：
Sanders2nd,CR;Schwonek,JP
通讯作者：
Schwonek,JP
影响因子：
56.9
作者：
KETCHEM, RR;HU, W;CROSS, TA
通讯作者：
CROSS, TA
来自非洲爪蟾卵母细胞膜的 Ca2 激活通道重构为平面双层。
DOI：
10.1073/pnas.81.16.5155
发表时间：
1984
影响因子：
11.1
作者：
Young,GP;Young,JD;Deshpande,AK;Goldstein,M;Koide,SS;Cohn,ZA
通讯作者：
Cohn,ZA
通过三维固态核磁共振光谱解析二维各向异性化学位移/异核偶极耦合粉末图案光谱。
DOI：
10.1006/jmrb.1996.0016
发表时间：
1996
期刊：
Journal of magnetic resonance. Series B.
影响因子：
--
作者：
Ramamoorthy,A;Gierasch,LM;Opella,SJ
通讯作者：
Opella,SJ