Department of Chemistry, University of Michigan, Ann Arbor, Michigan 48109, USA.
Biochemistry. 2010 Jul 13;49(27):5760-5. doi: 10.1021/bi100605e.
A variety of biologically active peptides exert their function through direct interactions with the lipid membrane of the cell. These surface interactions are generally transient and highly dynamic, making them hard to study. Here we have examined the feasibility of using solution phase (19)F nuclear magnetic resonance (NMR) to study peptide-membrane interactions. Using the antimicrobial peptide MSI-78 as a model system, we demonstrate that peptide binding to either small unilamellar vesicles (SUVs) or bicelles can readily be detected by simple one-dimensional (19)F NMR experiments with peptides labeled with l-4,4,4-trifluoroethylglycine. The (19)F chemical shift associated with the peptide-membrane complex is sensitive both to the position of the trifluoromethyl reporter group (whether in the hydrophobic face or positively charged face of the amphipathic peptide) and to the curvature of the lipid bilayer (whether the peptide is bound to SUVs or bicelles). (19)F spin echo experiments using the Carr-Purcell-Meiboom-Gill pulse sequence were used to measure the transverse relaxation (T(2)) of the nucleus and thereby examine the local mobility of the MSI-78 analogues bound to bicelles. The fluorine probe positioned in the hydrophobic face of the peptide relaxes at a rate that correlates with the tumbling of the bicelle, suggesting that it is relatively immobile, whereas the probe at the positively charged face relaxes more slowly, indicating this position is much more dynamic. These results are in accord with structural models of MSI-78 bound to lipids and point to the feasibility of using fluorine-labeled peptides to monitor peptide-membrane interactions in living cells.
多种具有生物活性的肽通过与细胞膜的直接相互作用发挥其功能。这些表面相互作用通常是短暂的和高度动态的，因此很难研究。在这里，我们研究了使用溶液相（19）F 核磁共振（NMR）研究肽-膜相互作用的可行性。我们使用抗菌肽 MSI-78 作为模型系统，证明了通过用 l-4,4,4-三氟乙基甘氨酸标记的肽进行简单的一维（19）F NMR 实验，可以容易地检测到肽与小单层囊泡（SUV）或双分子层囊泡的结合。与肽-膜复合物相关的（19）F 化学位移既对三氟甲基报告基团的位置（是否在两亲肽的疏水区或正电荷区）敏感，又对脂质双层的曲率（肽是结合到 SUV 还是双分子层囊泡）敏感。使用 Carr-Purcell-Meiboom-Gill 脉冲序列的（19）F 自旋回波实验用于测量核的横向弛豫（T（2）），从而检查结合到双分子层囊泡的 MSI-78 类似物的局部流动性。位于肽疏水区的氟探针以与双分子层囊泡翻滚相关的速率弛豫，表明其相对不移动，而位于正电荷区的探针弛豫较慢，表明该位置更动态。这些结果与 MSI-78 与脂质结合的结构模型一致，并表明使用氟标记的肽来监测活细胞中的肽-膜相互作用是可行的。
