Lipid-modulation of membrane insertion and refolding of the apoptotic inhibitor Bcl-xL.
Lipid-modulation of membrane insertion and refolding of the apoptotic inhibitor Bcl-xL.
复制标题
细胞凋亡抑制剂 Bcl-xL 的膜插入和重折叠的脂质调节。
DOI：
10.1016/j.bbapap.2019.04.006
复制
发表时间：
2019
期刊：
影响因子：
--
通讯作者：
Ladokhin,AlexeyS
中科院分区：
文献类型：
--
作者：
Vasquez-Montes,Victor;Vargas-Uribe,Mauricio;Pandey,NitinK;Rodnin,MykolaV;Langen,Ralf;Ladokhin,AlexeyS
Bcl-xL是凋亡调节因子Bcl-2家族的一员，负责抑制线粒体外膜的通透性，是一种很有前景的抗癌靶点。Bcl-xL存在于以下构象中，每个构象都被认为在抑制细胞凋亡中起作用：(a)可溶折叠构象，(b)膜锚定（通过其c端α8螺旋）形式，其保持与溶液中相同的折叠，(c)再折叠的膜插入构象，其结构数据可用。以往的研究证实，细胞内Bcl-xL处于可溶态和膜态之间的动态平衡状态，但在体内没有直接证据支持膜态的锚定构象或插入构象。在这项体外研究中，我们采用荧光和EPR光谱相结合的方法来表征Bcl-xL双层插入构象的结构特征以及脂质对其膜插入过渡的调节。结果表明，核心疏水螺旋α6不采用跨膜取向插入双分子层。这种插入破坏了Bcl-xL的包装，并从蛋白质结构的其余部分释放了调节n端BH4结构域（α1）。我们的数据表明，Bcl-xL的插入和重折叠都受到脂质成分的调节，这使得pKaof的插入达到生理ph值的阈值。我们假设与Bcl-xL双分子层插入相关的构象重排导致其切换到所谓的非规范凋亡抑制模式。这些结果表明，细胞凋亡前和凋亡过程中脂质组成的改变可能是调节线粒体外膜通透性的另一个因素。
Bcl-xL is a member of the Bcl-2 family of apoptotic regulators, responsible for inhibiting the permeabilization of the mitochondrial outer membrane, and a promising anti-cancer target. Bcl-xL exists in the following conformations, each believed to play a role in the inhibition of apoptosis: (a) a soluble folded conformation, (b) a membrane-anchored (by its C-terminal α8 helix) form, which retains the same fold as in solution and (c) refolded membrane-inserted conformations, for which no structural data are available. Previous studies established that in the cell Bcl-xL exists in a dynamic equilibrium between soluble and membranous states, however, no direct evidence exists in support of either anchored or inserted conformation of the membranous statein vivo. In thisin vitrostudy, we employed a combination of fluorescence and EPR spectroscopy to characterize structural features of the bilayer-inserted conformation of Bcl-xL and the lipid modulation of its membrane insertion transition. Our results indicate that the core hydrophobic helix α6 inserts into the bilayer without adopting a transmembrane orientation. This insertion disrupts the packing of Bcl-xL and releases the regulatory N-terminal BH4 domain (α1) from the rest of the protein structure. Our data demonstrate that both insertion and refolding of Bcl-xL are modulated by lipid composition, which brings the apparent pKaof insertion to the threshold of physiological pH. We hypothesize that conformational rearrangements associated with the bilayer insertion of Bcl-xL result in its switching to a so-called non-canonical mode of apoptotic inhibition. Presented results suggest that the alteration in lipid composition before and during apoptosis can serve as an additional factor regulating the permeabilization of the mitochondrial outer membrane.