Flip-flop is the rate-limiting step for transport of free fatty acids across lipid vesicle membranes.
Flip-flop is the rate-limiting step for transport of free fatty acids across lipid vesicle membranes.
复制标题
触发器是游离脂肪酸跨脂质囊泡膜运输的限速步骤。
DOI：
10.1021/bi901318a
复制
发表时间：
2009
期刊：
影响因子：
2.9
通讯作者：
Kleinfeld,AlanM
中科院分区：
文献类型：
--
作者：
Carley,AndrewN;Kleinfeld,AlanM
游离脂肪酸（FFA）在脂质双分子层膜上的转运机制仍然是一个有争议的话题。争论的焦点是转移的限速步骤是在膜上翻转还是解离进入水相。近年来，研究人员在脂质囊泡外叶中引入荧光素磷脂酰乙醇胺（FPE）来监测游离脂肪酸的分离。将FFA运输到含有FPE和pyranine的囊泡中，FPE和pyranine的荧光猝灭速率常数相同。由于观察到游离脂肪酸结合到外表面和在双分子层上翻转的时间没有差异，因此可以得出结论，游离比翻转慢。在这里，我们使用FPE和BSA来评估油酸与脂质囊泡的分离。在单独的含有吡嗪或adifab的囊泡中，我们评估了触发器。我们发现FPE和BSA转移方法产生的解离速率常数相等，比触发器快3 ~ 10倍。我们发现在同时含有FPE和pyranine的囊泡中，pyranine荧光不能与FPE荧光分离。在含有两种荧光团的囊泡中观察到的FPE和pyranine相同的速率常数反映了FPE荧光在pyranine激发和发射波长上的优势（20倍）。因为解离速率常数是flip-flop速率常数的3 - 10倍，所以flip-flop必须是FFA在脂质囊泡中运输的限速步骤。
The mechanism of transport of free fatty acids (FFA) across lipid bilayer membranes remains a subject of debate. The debate is whether the rate-limiting step for transport is flip-flop across the membrane or dissociation into the aqueous phase. Recently, a new method for assessing dissociation was described in which fluorescein phosphatidylethanolamine (FPE) introduced into the outer leaflet of lipid vesicles was used to monitor FFA dissociation. Transport of FFA into vesicles containing both FPE in the outer leaflet and pyranine trapped in the inside aqueous phase revealed identical rate constants for quenching of FPE and pyranine fluorescence. Because no difference was observed in the time for FFA binding to the outer surface and flip-flop across the bilayer, it was concluded that dissociation was slower than flip-flop. Here, we used FPE and BSA to assess dissociation of oleate from lipid vesicles. In separate pyranine- or ADIFAB-containing vesicles, we assessed flip-flop. We found that the FPE and BSA transfer methods yielded equivalent rate constants for dissociation, which were 3−10-fold faster than that of flip-flop. We found that in vesicles containing both FPE and pyranine, pyranine fluorescence cannot be separated from FPE fluorescence. The identical rate constants for FPE and pyranine observed with vesicles containing both fluorophores reflected the dominance (20-fold) of FPE fluorescence at pyranine excitation and emission wavelengths. Because the dissociation rate constants are 3−10 times faster than the rate constants for flip-flop, flip-flop must be the rate-limiting step for the transport of FFA across lipid vesicles.