硫化醣鍵結物在調節毒素與生長因子的內吞作用上的角色

Endocytosis of toxins and growth factors: A role of sulfo-glycoconjugates in tuning specificity and capacity

劉益安(Yi-An Liou)

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(2007年)

摘要

細胞膜表面是巨分子經由內吞作用進入細胞的一道關卡而且膜上具有許多的接受器與硫化乙醯肝素蛋白多醣。硫化乙醯肝素蛋白多醣在協調哺乳類的生理機能上(例如協調消化、內分泌、神經系統等)扮演重要的角色。本文在描述硫化乙醯肝素蛋白多醣在貨物進入細胞的生物功能。肝癌衍生生長因子的HATH區域和心臟毒素被用來研究硫化乙醯肝素蛋白多醣在內吞作用上的功能。肝素酶的作用在於切除硫化乙醯肝素的硫化乙醯肝素進而干擾高濃度的HATH區域進入細胞而競爭劑能夠抑制低濃度的HATH區域經由接受器進入細胞。在本實驗中，我們證明了HATH區域進入細胞的量的增加是由於硫化乙醯肝素蛋白多醣的影響所致，並且直接證明了HATH區域進入細胞的早期和晚期分別是受到接受器與硫化乙醯肝素蛋白多醣的影響。在心臟毒素A2與A4的蛋白質的序列中，一個胺基酸的差異能引起在細胞素性與進入細胞的量上的多樣性。心臟毒素A2與A4分別對低硫酸化與高硫酸化的硫化乙醯肝素蛋白多醣具有專一性並且其進入細胞的途徑不是經由包涵素引導與胞膜窖引導之內吞作用而這兩個途徑被報導與接受器引導之內吞作用有關。沒有證據顯示硫化乙醯肝素蛋白多醣引導的內吞作用與接受器有關，而心臟毒素A4在以往的研究顯示與富有磷脂醯肌醇(蛋白)聚糖的硫化乙醯肝素蛋白多醣會經由同一條路徑進入細胞。雖然心臟毒素A2與A4可能會經由硫化乙醯肝素蛋白多醣調控的途徑進入細胞但是兩者會與不同的硫化乙醯肝素蛋白多醣產生作用，這樣的選擇性或許能夠解釋心臟毒素只對特定的細胞產生較大毒性的原因。

關鍵字

內吞作用 ； 心臟毒素 ； 生長因子

並列摘要

The plasma membrane is a barrier for endocytosis of macromolecules. There are many receptors and heparan sulfate proteoglycans (HSPG) at cell surface. HSPG plays a very important role in tuning the mammalian physiology including digestive, endocrine, nervous, and other systems. This study investigated the biological role of HSPG at internalization of cargos, using HATH domain of hepatoma-derived growth factor (HDGF) and cardiotoxins (CTX) A2 / A4 as two examples. The treatment of heparinase interfered the internalization at high dose of HATH domain and competition against low dose HATH domain inhibited the receptor mediated endocytosis pathway. In this study we demonstrate the enhancement of internalization of HATH domain was caused by HSPG. The early and late stages of internalization of HATH domain were through receptor and HSPG mediated endocytosis, respectively. One residue difference resulted in considerable difference in the amount of internalization between CTX A2 and A4. CTX A2 and A4 were specific to low or high sulfate HSPG and does not internalize through clathrin or caveolae mediated endocytosis. These two pathways were concern with receptor mediated endocytosis. HSPG mediated endocytosis was not via receptor mediated pathway and CTX A4 was via glypican enriched HSPG pathway. Although the CTX A2 and A4 maybe internalize through HSPG mediated endocytosis but they may interact with the different domain HSPG.

並列關鍵字

Endocytosis ； CTX ； cardiotoxin ； growth factor ； HSPG

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