Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, Arkansas 72701, United States.
Biochemistry. 2012 Mar 13;51(10):2044-53. doi: 10.1021/bi201732e. Epub 2012 Mar 5.
Synthetic model peptides have proven useful for examining fundamental peptide-lipid interactions. A frequently employed peptide design consists of a hydrophobic core of Leu-Ala residues with polar or aromatic amino acids flanking each side at the interfacial positions, which serve to "anchor" a specific transmembrane orientation. For example, WALP family peptides (acetyl-GWW(LA)(n)LWWA-[ethanol]amide), anchored by four Trp residues, have received particular attention in both experimental and theoretical studies. A recent modification proved successful in reducing the number of Trp anchors to only one near each end of the peptide. The resulting GWALP23 (acetyl-GGALW(5)(LA)(6)LW(19)LAGA-[ethanol]amide) displays reduced dynamics and greater sensitivity to lipid-peptide hydrophobic mismatch than traditional WALP peptides. We have further modified GWALP23 to incorporate a single tyrosine, replacing W(5) with Y(5). The resulting peptide, Y(5)GWALP23 (acetyl-GGALY(5)(LA)(6)LW(19)LAGA-amide), has a single Trp residue that is sensitive to fluorescence experiments. By incorporating specific (2)H and (15)N labels in the core sequence of Y(5)GWALP23, we were able to use solid-state NMR spectroscopy to examine the peptide orientation in hydrated lipid bilayer membranes. The peptide orients well in membranes and gives well-defined (2)H quadrupolar splittings and (15)N/(1)H dipolar couplings throughout the core helical sequence between the aromatic residues. The substitution of Y(5) for W(5) has remarkably little influence on the tilt or dynamics of GWALP23 in bilayer membranes of the phospholipids DOPC, DMPC, or DLPC. A second analogue of the peptide with one Trp and two Tyr anchors, Y(4,5)GWALP23, is generally less responsive to the bilayer thickness and exhibits lower apparent tilt angles with evidence of more extensive dynamics. In general, the peptide behavior with multiple Tyr anchors appears to be quite similar to the situation when multiple Trp anchors are present, as in the original WALP series of model peptides.
合成模型肽已被证明可用于研究基本的肽-脂相互作用。一种常用的肽设计由亮氨酸-丙氨酸残基组成的疏水区核心组成，每个界面位置的两侧都有极性或芳香族氨基酸，这些氨基酸用于“锚定”特定的跨膜取向。例如，WALP 家族肽（乙酰基-GWW(LA)(n)LWWA-[乙醇]酰胺），由四个色氨酸残基锚定，在实验和理论研究中都受到特别关注。最近的一项修改成功地将色氨酸锚的数量减少到肽的每端仅一个。由此产生的 GWALP23（乙酰基-GGALW(5)(LA)(6)LW(19)LAGA-[乙醇]酰胺）显示出降低的动力学和对脂质-肽疏水性失配的更高敏感性，比传统的 WALP 肽。我们进一步修改了 GWALP23，在其中引入一个酪氨酸，用 Y(5)取代 W(5)。由此产生的肽，Y(5)GWALP23（乙酰基-GGALY(5)(LA)(6)LW(19)LAGA-酰胺），只有一个色氨酸残基对荧光实验敏感。通过在 Y(5)GWALP23 的核心序列中掺入特定的 (2)H 和 (15)N 标记，我们能够使用固态 NMR 光谱法研究水合脂质双层膜中肽的取向。肽在膜中定向良好，并在芳香族残基之间的核心螺旋序列中给出清晰的 (2)H 四极分裂和 (15)N/(1)H 偶极耦合。用 Y(5)取代 W(5)对 GWALP23 在 DOPC、DMPC 或 DLPC 磷脂双层膜中的倾斜或动力学的影响极小。肽的第二个类似物，一个色氨酸和两个酪氨酸锚，Y(4,5)GWALP23，对双层厚度的响应通常较小，并且表现出较低的表观倾斜角，并显示出更广泛的动力学证据。一般来说，具有多个 Tyr 锚的肽的行为似乎与存在多个 Trp 锚的情况非常相似，就像原始 WALP 系列模型肽一样。
