以原子力顯微鏡探討膽固醇及心臟毒素A3對腦硫脂人造細胞膜的作用

AFM studies of Cholesterol and Cardiotoxin-A3 Effects on Sulfatide-POPC Model Membranes

陳弘銘(Chen, Hung Ming)

指導教授 ： 吳文桂

國立清華大學/生命科學院/生物資訊與結構生物研究所/碩士(2010年)

https://doi.org/10.6843/NTHU.2010.00227

摘要

摘要

過去的研究發現，膽固醇與腦硫脂對細胞膜的狀態以及心臟毒素A3的作用有很大的影響。因此在本論文中，首先利用原子力顯微鏡闡明膽固醇對細胞膜狀態的影響。接著透過原子力顯微鏡以及共軛焦顯微鏡觀察量測，以確定腦硫脂與細胞膜的作用區域。結果發現膽固醇對腦硫脂-POPC雙層膜造成的效果與過去文獻中所記載的其他脂質相符合；透過原子力顯微鏡以及共軛焦顯微鏡，我們也發現心臟毒素A3的作用區域確實是在腦硫脂上，且原子力顯微鏡的證據顯示，心臟毒素A3有可能是透過腦硫脂的邊緣區域結合於膜上後，再進一步插入膜內，此結果能進一步證明過去本實驗室所推論出的心臟毒素-腦硫脂作用模型。藉此使我們對心臟毒素的作用機制有更確實的了解。

關鍵字

心臟毒素A3 ； 原子力顯微鏡 ； 腦硫脂

並列摘要

Abstract

It has recently been shown that cholesterol and sulfatides plays important role in cell membrane status and CTX-A3 interactions. In order to understand the relationship between cholesterol, model membrane and CTX-A3, we employed atomic force microscopy to study cholesterol and model membrane relationship first. We also applied confocal microscope to study the interaction between CTX-A3 and model membrane. Our results suggest that cholesterol decrease the height of sulfatide domains on model membrane, also decrease the average membrane thickness of Sulfatide- POPC membrane. We also found CTX-A3 interacts with sulfatide domains. According to statistic results of AFM images, we found the initial interaction of CTX-A3 and sulfatide-POPC membrane might happen at the edge of sulfatide domains. Which means CTX-A3 might attach to the edge of sulfatide domain, and then CTX-A3 could disturb the dynamics of the edge lipids. Finally CTX-A3 could interact with other CTX-A3 to form complex or insert to membrane to induce following changes. This confirms our labs model again and gives us more understanding of that how CTX-A3 could interact with cell membranes.

參考文獻

41. 林志成，林世明 (2007) 奈米量測技術－原子力顯微鏡在生物分子上之應用, 化學, 第六十七卷, 83-91頁

1. Pinto-Alphandary H., Andremont A.(2000) "Targeted delivery of antibiotics using liposomes and nanoparticles: research and applications." Int. J. Antimicrob. Agents. 13(3), 155-168.

2. Reviakine I., Brisson A., et al.(2000) "Formation of supported phosolipid bilayers from unilamellar vesicles investigated by atomic force microscopy", Langmuir, 16,1806-1815

6.Jiang, M.S., Fletcher, et al. (1989) "Factors influencing the hemolysis of human erythrocytes by cardiotoxins from Naja naja kaouthia and Naja naja atra venoms and a phospholipase A2 with cardiotoxin-like activities from Bungarus fasciatus venom" Toxicon 27, 247-257.

7. Kini, R.M., and Evans, H.J. et al (1989) "Role of Cationic Residues in Cytolytic Activity: Modification of Lysine Residues in the Cardiotoxin from Naja nigricollis Venom and Correlation between Cytolytic and Antiplatelet Activity" Biochemistry 28, 9209-9215.