Kurrle A, Rieber P, Sackmann E
Physik Department, Technische Universität München, Garching, Germany.
Biochemistry. 1990 Sep 11;29(36):8274-82. doi: 10.1021/bi00488a011.
We studied the interaction of transferrin receptors (of cell line Molt-4) with mixed model membranes as a function of lipid chain length (phospholipids with C14:0 and C18:1 hydrocarbon chains) and of the surface charge of the membrane using mixtures of C14:0 lecithin (DMPC) with C14:0 phosphatidylglycerol (DMPG) and C14:0 phosphatidylserine (DMPS). Spontaneous self-assembly of receptors and lipids was achieved by freeze-thaw cycles of a codispersion of mixed vesicles and receptors in buffer and subsequent separation of receptor-loaded and receptor-free vesicles by density gradient centrifugation. Information on specific lipid/protein interaction mechanisms was obtained by evaluation of protein-induced shifts of phase boundaries of lipid mixtures by calorimetry and by FTIR spectroscopy of partially deuterated lipid mixtures. The important role (1) of minimizing the elastic forces caused by the mismatch of the lengths of hydrophobic cores of the protein (lp) and the bilayer (lL) and (2) of the electrostatic coupling of protein head groups with the charged membrane/water interface for the lipid/protein self-assembly is established. The electrostatic interaction energy per receptor is about 10(3) kBT (by coupling to about 1000 charged lipids) which is sufficient to overcompensate the elastic energy associated with a mismatch of lp - lL approximately 1.0 nm. The maximum receptor concentration incorporated was measured as a function of membrane surface charge and lipid chain length. The maximum receptor molar fraction varied from xpmax = 5 x 10(-5) for DMPC to xpmax = 4 x 10(-4) for 1:1 DMPC/DMPG; moreover xpmax is higher for DMPS than for DMPG as charged component. For the long-chain lipids, xpmax is higher for a 9:1 DEPE/DEPC mixture [(4.2-9) x 10(-4)] than for pure DEPC (ca. 3.5 x 10(-4)). By decomposition of reconstituted receptors with proteases, we demonstrated the homogeneous orientation of the receptor with its extracellular head group pointing to the convex side of the vesicles.(ABSTRACT TRUNCATED AT 250 WORDS)
我们研究了（细胞系Molt-4的）转铁蛋白受体与混合模型膜之间的相互作用，该相互作用是脂质链长度（具有C14:0和C18:1烃链的磷脂）以及膜表面电荷的函数，其中膜表面电荷通过C14:0卵磷脂（DMPC）与C14:0磷脂酰甘油（DMPG）和C14:0磷脂酰丝氨酸（DMPS）的混合物来调控。受体和脂质的自发自组装通过缓冲液中混合囊泡与受体的共分散液的冻融循环，以及随后通过密度梯度离心分离载有受体和无受体的囊泡来实现。通过量热法评估蛋白质诱导的脂质混合物相界移动以及对部分氘代脂质混合物进行傅里叶变换红外光谱（FTIR）分析，获得了关于特定脂质/蛋白质相互作用机制的信息。确定了（1）最小化由蛋白质疏水核心长度（lp）与双层膜长度（lL）不匹配引起的弹力，以及（2）蛋白质头部基团与带电膜/水界面的静电耦合在脂质/蛋白质自组装中的重要作用。每个受体的静电相互作用能约为10(3) kBT（通过与约1000个带电脂质耦合），这足以过度补偿与lp - lL约1.0 nm的不匹配相关的弹性能。测量了掺入的最大受体浓度作为膜表面电荷和脂质链长度的函数。最大受体摩尔分数从DMPC的xpmax = 5 x 10(-5)变化到1:1 DMPC/DMPG的xpmax = 4 x 10(-4)；此外，作为带电成分，DMPS的xpmax高于DMPG。对于长链脂质，9:1 DEPE/DEPC混合物的xpmax（(4.2 - 9) x 10(-4)）高于纯DEPC（约3.5 x 10(-4)）。通过用蛋白酶分解重组受体，我们证明了受体的均匀取向，其细胞外头部基团指向囊泡的凸面。（摘要截断于250字）
