鹼性多肽鏈如何進入細胞一直是個具有爭議性且複雜的議題，但具有重要生物意義。台灣眼鏡蛇毒蛋白裡一個主要的成分為心臟毒素，各心臟毒蛋白(cardiotoxin)之間為一個結構相似的β-板狀(β-sheet polypeptides)且帶有正電荷的蛋白，使被咬傷者的心肌收縮而停止跳動，但是對於其進入細胞的詳細機制並不是很了解。(一)在過去研究中，心臟毒素中A2、A4會在進入細胞後並進一步的產生毒性，而在心臟毒素A2、A4的胺基酸的序列中之間有一個胺基酸的差異且心臟毒素A2、A4之間對於硫化乙醯肝素蛋白多醣 (HSPG)識別具有差異性，因此在本研究想要去探討心臟毒素A2，A4進入細胞的機制。結果顯示，A2、A4兩者具有相同程度的巨胞飲作用(Macropinocytosis)以及胞膜窖引導之內吞作用(Caveolae-mediated endpcytosis)，但卻有不同程度由包涵素引導的內吞作用(Clathrin-mediated endocytosis)，因此一個胺基酸在蛋白上的差異所能引起細胞胞吞反應的差異性。(二)心臟毒素A3在過去研究中，由於發現具有形成孔洞的能力，會造成孔洞的蛋白進入細胞的機制與細胞膜修復有關聯，我們發現心臟毒素A3會在細胞表面上造成不具選擇性的孔洞，在外部有鈣的環境下，隨著鈣離子經由心臟毒素A3產生的孔洞流入而引導細胞膜修復，然而在細胞外部沒有鈣離子的情況下，心臟毒素A3更容易進入細胞且聚集在粒線體。因此，除了鈣離子之外，膽固醇亦會調控心臟毒素的A3進入細胞，影響心臟毒素A3進入細胞的量和進入細胞的位置。上述的結果顯示鹼性多肽鏈對於硫化乙醯肝素蛋白多醣和膽固醇可能有不同的敏感性，造成對於細胞反應有所不同，這顯示蛋白質進入細胞的機制有多重的調控，而且對細胞膜的醣胺素及脂質有高靈敏度。

How basic polypeptide endocytosis got into living cells was a controversial and complex issue but with significant biological function. Cardiotoxins (CTXs), a major component of snake venom from Taiwan cobra Naja atra, are structurally homologues beta-sheet basic polypeptides that cause systolic cardiac arrest of the bitten victim with unknown internalization mechanism. (1) Previous study showed that CTX A2, A3 and A4 would get into cell and further exhibited toxicities; moreover, comparing amino acid between CTX A2 and A4, there are one amino acid difference. CTX A2 and A4 were specific to low or high sulfate heparan sulfate proteoglycans (HSPG), respectively. It is important to know what the endocytosis route of CTX A2, A3 and A4 is. According to these results, CTX A2 and A4 internalized via macropinocytosis and caveolae-mediated endocytosis at the same level but not for clathrin-mediated endocytosis. Therefore, one amino acid difference on protein caused different cell internalization responses in cells. (2) In the previous study, CTX A3 will cause pore-formation. Several researches show the correlation between membrane repair and internalization. We found that CTX A3 formed none-specific pore on cell membrane based on the result. In the presence of calcium, calcium influx from CTX A3-inducing pore triggered membrane repair process. However, in the absence of calcium, CTX A3 got into cell and targeted to mitochondria more obvious compared with in the presence of calcium environment. Besides calcium effect, cholesterol could also regulate CTX A3 internalization and localization. According to these data, basic polypeptide probably with different sensitive to HSPG and cholesterol caused different cell response. This indicated that protein internalization was regulated by multiple mechanisms and with high sensitive to HSPG and cholesterol of cell membrane.

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