Immobilizing single lipid and channel molecules in artificial lipid bilayers with annexin A5
Immobilizing single lipid and channel molecules in artificial lipid bilayers with annexin A5
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DOI：
10.1021/la0535025
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发表时间：
2006-07-04
期刊：
影响因子：
3.9
通讯作者：
Ide, Toru
中科院分区：
文献类型：
--
作者：
Ichikawa, Takehiko;Aoki, Takaaki;Ide, Toru
研究了膜联蛋白A5对单分子脂质和单分子蛋白质在人工脂双层膜中横向扩散的影响。膜联蛋白A5是膜联蛋白超家族的成员，其以Ca 2+依赖性方式优先结合阴离子磷脂。在这份报告中，我们能够直接监测单BODIPY 1，2-双十六烷酰基-sn-甘油基-3-磷酸乙醇胺（DHPE）和兰尼碱受体2型（RyR 2）标记的Cy 5分子的脂质双层含有磷脂酰丝氨酸（PS）通过使用荧光显微镜。在各种膜联蛋白A5浓度下计算扩散系数。在不存在膜联蛋白A5的情况下，BODIPY-DHPE和Cy 5-RyR 2的扩散系数分别为4.81 x 10（-8）cm（2）/s和2.13 x 10（-8）cm（2）/s。在1 AM膜联蛋白A5存在下，BODIPY-DHPE和Cy 5-RyR 2的扩散系数分别为2.2 × 10（-10）cm（2）/s和9.5 × 10（-11）cm（2）/s。总体而言，1 μ M的膜联蛋白A5足以诱导横向扩散系数降低200倍。此外，我们进行了电生理检查，并确定膜联蛋白A5对RyR 2的功能几乎没有影响。这意味着当成像和分析RyR 2时，膜联蛋白A5可用于在脂质双层中包裹RyR 2。
The effects of annexin A5 on the lateral diffusion of single-molecule lipids and single-molecule proteins were studied in an artificial lipid bilayer membrane. Annexin A5 is a member of the annexin superfamily, which binds preferentially to anionic phospholipids in a Ca2+-dependent manner. In this report, we were able to directly monitor single BODIPY 1,2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine (DHPE) and ryanodine receptor type 2 (RyR2) labeled with Cy5 molecules in lipid bilayers containing phosphatidylserine (PS) by using fluorescence microscopy. The diffusion coefficients were calculated at various annexin A5 concentrations. The diffusion coefficients of BODIPY-DHPE and Cy5-RyR2 in the absence of annexin A5 were 4.81 x 10(-8) cm(2)/s and 2.13 x 10(-8) cm(2)/s, respectively. In the presence of 1 AM annexin A5, the diffusion coefficients of BODIPY-DHPE and Cy5-RyR2 were 2.2 x 10(-10) cm(2)/s and 9.5 x 10(-11) cm(2)/s, respectively. Overall, 1 mu M of annexin A5 was sufficient to induce a 200-fold decrease in the lateral diffusion coefficient. Additionally, we performed electrophysiological examinations and determined that annexin A5 has little effect on the function of RyR2. This means that annexin A5 can be used to immobilize RyR2 in a lipid bilayer when imaging and analyzing RyR2.