Abstract
中文摘要 有毒之海洋生物，往往含有豐富的化學物質或毒素，這此物 質常具有抗腫瘤、抗癌、抗病毒、抗菌、抗發炎等生理活性、具有被開發 作為天然藥物資源的潛力。 scratcher 為芋螺所分泌的毒素之一，它 的作用接受體尚未被證實，主要症狀為使實驗動物產生搔癢現象。本身為 一段含有 29 氨基酸的 peptide ，其中包括4 個半胱氨酸，可形成2個雙 硫鍵，此外，也包括一個轉譯後修飾的氨基酸--carboxyglutamate(Gla) 且碳端出現的醯胺化(amidation)現象。 實驗上，我們利用合成方式 製造這一段peptide ，並利用高效液相層析法加以純化。經由質譜鑑定分 子量無誤，確定為還原態scratcher 。接著在以Tris buffer 為鹼性環境 下進行大氣氧化。在氧化過程中我們發現幾乎以某種特定雙硫鍵鍵結方式 為主(可能為[12-25;16-27]或[12-27;16-25])。且在CD實驗中發現將近30 ％螺旋性。此外，我們亦用不同濃度的GudHCl 去探討Scratcher之其他他 特性。最後分別在以水及氘為溶液進行不同的核磁共振實驗 (TOCSY、 NOESY及 DQF-COSY) ，並根據NOESY光譜所得到的距離限制，配合分子模 擬 軟體加以分析計算，以解出最可能的結構。而最終結果我們發現在 Phe7至Ile20形成一穩定之螺旋結構。 Abstract Many marine organisms usually secrete chemical compounds and toxins,which posses antitumor, antiviral, antibacterial, antinflammatoryactivities. The toxins are therefore candidates for drugs. Scratcher is a kind of venom secreted by cone snail. Thebiochemical property of scratcher is still uncertain, but the amino acidsequence and the symptoms of scratch toxicity in animal test are known.Scratcher is a small peptide, composed of 29 amino acid residues, andcontains 4 cysteines, forming two disulfide bonds. Scratcher also containsa carboxyglutamic acid and amidation at C-terminal, which are rare post-translational modifications. We synthesized and purified scratcher and checked the molecularweight by Mass spectroscopy. After oxidation reactions, we found onemajor product which may be some disulfide bond patterns ([12-27;16-25]or [12-25;16-27]). Our CD data indicated that the structure of scratcher iscomposed of 30% helicity. We also determined the solution structure of scratcher in water, using a combination of NMR data and distancegeometry calculation. The NMR data corroborates our CD data: we observed a characteristic a-helical conformation in the sequence region ofPhe7-Ile20.