Matsubara Teruhiko, IIjima Kazutoshi, Kojima Takahiro, Hirai Miwa, Miyamoto Erika, Sato Toshinori
Department of Biosciences and Informatics, Keio University, 3-14-1 Hiyoshi, Kouhoku-ku, Yokohama 223-8522, Japan.
Langmuir. 2021 Jan 19;37(2):646-654. doi: 10.1021/acs.langmuir.0c02387. Epub 2021 Jan 5.
The specific features of the lateral distribution of gangliosides play key roles in cell-cell communications and the onset of various diseases related to the plasma membrane. We herein demonstrated that an artificial peptide identified from a phage-displayed library is available as a molecular probe for specific ganglioside nanoclustering sites in caveolae/membrane rafts on the cell surface. Atomic force microscopy studies indicated that the peptide specifically binds to the highly enriched monosialoganglioside GM1 nanodomains of reconstituted lipid bilayers composed of GM1, sphingomyelin, cholesterol, and unsaturated phospholipids. The ganglioside-containing area recognized by the peptide on the surface of PC12 cells was part of the area recognized by the cholera toxin B subunit, which has high affinity for GM1. Furthermore, the peptide bound to the cell surface after a treatment with methyl-β-cyclodextrin (MβCD), which disrupts membrane rafts by removing cholesterol. The present results indicate that there are heterogeneous ganglioside clusters with different ganglioside densities in caveolae/membrane rafts, and the peptidyl probe selectively recognizes the high-density ganglioside nanodomain that resists the MβCD treatment. This peptidyl probe will be useful for obtaining information on the lipid organization of the cell membrane and will help clarify the mechanisms by which the lateral distribution of gangliosides affects biological functions and the onset of diseases.
神经节苷脂侧向分布的特定特征在细胞间通讯以及与质膜相关的各种疾病的发生中起着关键作用。我们在此证明，从噬菌体展示文库中鉴定出的一种人工肽可作为一种分子探针，用于检测细胞表面小窝/膜筏中特定的神经节苷脂纳米簇位点。原子力显微镜研究表明，该肽特异性结合由GM1、鞘磷脂、胆固醇和不饱和磷脂组成的重组脂质双层中高度富集的单唾液酸神经节苷脂GM1纳米结构域。该肽在PC12细胞表面识别的含神经节苷脂区域是霍乱毒素B亚基识别区域的一部分，霍乱毒素B亚基对GM1具有高亲和力。此外，在用甲基-β-环糊精（MβCD）处理后，该肽与细胞表面结合，MβCD通过去除胆固醇破坏膜筏。目前的结果表明，在小窝/膜筏中存在具有不同神经节苷脂密度的异质性神经节苷脂簇，并且该肽基探针选择性地识别抗MβCD处理的高密度神经节苷脂纳米结构域。这种肽基探针将有助于获取有关细胞膜脂质组织的信息，并有助于阐明神经节苷脂侧向分布影响生物学功能和疾病发生的机制。
