De Kroon A I, Soekarjo M W, De Gier J, De Kruijff B
Centre for Biomembranes and Lipid Enzymology, University of Utrecht, The Netherlands.
Biochemistry. 1990 Sep 11;29(36):8229-40. doi: 10.1021/bi00488a006.
The interaction of interrelated model peptides with model membranes has been studied by techniques based on tryptophan fluorescence. The peptides used are derivatives of the sequence H-Ala-Met-Leu-Trp-Ala-OH, which was designed for this purpose. Several modifications yielded a set of 13 penta- and hexapeptides varying in net charge, hydrophobicity, charge distribution, and the intramolecular position of the tryptophan residue with respect to the charge(s). The affinity of these peptides for small unilamellar vesicles (SUV) consisting of zwitterionic egg phosphatidylcholine (eggPC) and negatively charged beef heart cardiolipin (bhCL) has been investigated in a comparative way. The criteria for affinity comprise (1) intrinsic fluorescence changes upon titration of the peptides with the lipid vesicles, (2) reduced accessibility of the peptides to aqueous quenchers of tryptophan fluorescence (I- and acrylamide) in the presence of lipid, and (3) exposure to membrane-incorporated fluorescence quenchers, brominated phosphatidylcholines (BrPC). Application of BrPC brominated at different positions along the acyl chains provided information on the membrane topology of the peptides. With respect to the extent of affinity for zwitterionic membranes, the overall hydrophobicity of the peptides is the main determinant. A comparison of the affinity for PC of equally hydrophobic peptides carrying either a single positive or negative charge reveals preferential interaction of the cationic peptide. Both hydrophobic and electrostatic interactions determine the affinity of positively charged mono- and divalent peptides for CL vesicles. The distribution of the charged moieties in divalent positively charged peptides, either both at one end of the molecule or one at each end, has little influence on the affinity of these peptides for CL but does affect the extent of exposure to BrPC. Upon decreasing the surface charge density of the vesicles by diluting CL with increasing amounts of PC, both types of peptides show different behavior. The position of the tryptophan relative to the charged moiety in the peptide molecule is shown to affect the fluorescent properties upon interaction with vesicles. Concerning the membrane topology, all peptides adopt a localization near the membrane surface, with the neutral peptides inserting slightly deeper into the bilayer than the charged peptides. The results allow a comparative analysis of the factors determining the extents and modes of lipid-model peptide interaction; in addition, the validity of the methods applied is discussed.
通过基于色氨酸荧光的技术研究了相互关联的模型肽与模型膜的相互作用。所使用的肽是为此目的设计的序列H-Ala-Met-Leu-Trp-Ala-OH的衍生物。几种修饰产生了一组13种五肽和六肽，它们在净电荷、疏水性、电荷分布以及色氨酸残基相对于电荷的分子内位置方面有所不同。以比较的方式研究了这些肽对由两性离子鸡蛋磷脂酰胆碱（eggPC）和带负电荷的牛心磷脂（bhCL）组成的小单层囊泡（SUV）的亲和力。亲和力的标准包括：（1）用脂质囊泡滴定肽时的固有荧光变化；（2）在脂质存在下，肽对色氨酸荧光的水性猝灭剂（I-和丙烯酰胺）的可及性降低；（3）暴露于膜结合的荧光猝灭剂溴化磷脂酰胆碱（BrPC）。沿酰基链在不同位置溴化的BrPC的应用提供了有关肽的膜拓扑结构的信息。就对两性离子膜的亲和力程度而言，肽的整体疏水性是主要决定因素。对带有单个正电荷或负电荷的同等疏水性肽对PC的亲和力进行比较，发现阳离子肽具有优先相互作用。疏水性和静电相互作用都决定了带正电荷的单价和二价肽对CL囊泡的亲和力。二价带正电荷肽中带电部分的分布，无论是在分子的一端还是两端各一个，对这些肽对CL的亲和力影响很小，但确实会影响暴露于BrPC的程度。通过用越来越多的PC稀释CL来降低囊泡的表面电荷密度时，两种类型的肽表现出不同的行为。结果表明，肽分子中色氨酸相对于带电部分的位置在与囊泡相互作用时会影响荧光特性。关于膜拓扑结构，所有肽都定位于膜表面附近，中性肽比带电肽稍微更深地插入双层中。这些结果允许对决定脂质-模型肽相互作用程度和模式的因素进行比较分析；此外，还讨论了所应用方法的有效性。
