Membrane activity of the phospholipase C-δ1 pleckstrin homology (PH) domain
Membrane activity of the phospholipase C-δ1 pleckstrin homology (PH) domain
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DOI：
10.1042/bj20041721
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发表时间：
2005-07-15
影响因子：
4.1
通讯作者：
Burger, KN
中科院分区：
文献类型：
--
作者：
Flesch, FM;Yu, JW;Burger, KN
PH-PLC8，[PLC Delta(1)(磷脂酶C-Delta(1))的PH结构域(Pleckstrin Homology Domaines)]是最典型的磷脂酰肌醇结合结构域之一。PH-PLC Delta(1)与PtdIns(4，5)P-2的头基具有高度的特异性，但对其界面性质知之甚少。在本研究中，我们发现PH-PLC Delta(1)也是膜活性的，并且在生理(相当于双分子层)的表面压力下可以显著地插入含PtdIns(4，5)P-2的单层膜。然而，这种膜活性似乎涉及不同于靶向PH-PLC Delta(1)到PtdIns(4，5)P-2头基的相互作用。而大部分PtdIns(4，5)P-2结合的PH-PLC Delta(1)可以通过加入过量的可溶性头基[Ins(1，4，5)P-3]来取代，而PH-PLC Delta(1)的膜活性不能被取代。PH-PLC Delta(1)与其他磷脂酰肌醇结合区的不同之处在于，它的膜插入不需要占据磷脂酰肌醇结合部位。当磷脂酰肌醇结合位点发生突变时，仍然存在显著的单层插入，并且PH-PLC Delta(1)可以插入到根本不含PtdIns(4，5)P-2的单层中。我们的结果表明，在PtdIns(4，5)P-2或其他酸性磷脂的介导下，PH-PLC Delta(1)的可逆膜结合发生在没有膜插入的情况下。PH结构域在膜表面的积累提高了插入效率，但对插入的程度没有显著影响，而磷脂酰乙醇胺和胆固醇的存在促进了插入的程度。这是分离的PH结构域中膜活性的第一次报道，对于理解这种常见类型的结构域的膜靶向性具有重要意义。
PH-PLC8, [the PH domain (pleckstrin homology domain) of PLC delta(1) (phospholipase C-delta(1))] is among the best-characterized phosphoinositide-binding domains. PH-PLC delta(1) binds with high specificity to the headgroup of PtdIns(4,5)P-2, but little is known about its interfacial properties. In the present study, we show that PH-PLC delta(1) is also membrane-active and can insert significantly into PtdIns(4,5)P-2-containing monolayers at physiological (bilayer-equivalent) surface pressures. However, this membrane activity appears to involve interactions distinct from those that target PH-PLC delta(1) to the PtdIns(4,5)P-2 headgroup. Whereas the majority of PtdIns(4,5)P-2-bound PH-PLC delta(1) can be displaced by adding excess of soluble headgroup [Ins(1,4,5)P-3], membrane activity of PH-PLC delta(1) cannot. PH-PLC delta(1) differs from other phosphoinositide-binding domains in that its membrane insertion does not require that the phosphoinositide-binding site be occupied. Significant monolayer insertion remains when the phosphoinositide-binding site is mutated, and PH-PLC delta(1) can insert into monolayers that contain no PtdIns(4,5)P-2 at all. Our results suggest a model in which reversible membrane binding of PH-PLC delta(1), mediated by PtdIns(4,5)P-2 or other acidic phospholipids, occurs without membrane insertion. Accumulation of the PH domain at the membrane surface enhances the efficiency of insertion, but does not significantly affect its extent, whereas the presence of phosphatidylethanolamine and cholesterol in the lipid mixture promotes the extent of insertion. This is the first report of membrane activity in an isolated PH domain and has implications for understanding the membrane targeting by this common type of domain.