膠原蛋白模擬胜肽與硼烷共軛化合物的製備及雙硫鍵與cation-π作用力對膠原蛋白異源三股螺旋摺疊探討

膠原蛋白是哺乳動物中含量最多的蛋白質，也是人體內非常重要的蛋白質。膠原蛋白因為具有高度的生物相容性、低免疫性、生物降解性，已被廣泛的運用在生醫材料上。本研究的第一部分中，我們將硼烷分子 (boroncage)修飾到膠原蛋白模擬胜肽(POG)7上；硼烷分子是目前中子捕捉療法的其中一環，當硼原子接受到中子時會放出熱能進而殺死癌細胞，此研究期望利用膠原蛋白模擬胜肽的生物相容性，使此硼烷分子可以進入到Hela癌症細胞內。實驗結果顯示修飾上硼烷分子的(POG)7的熱穩定性僅有些微下降，且在細胞胞吞實驗中經過ICP- MASS的檢測發現細胞中有硼原子的訊號，所以此類膠原蛋白模擬胜肽具有運送硼進入細胞的潛在應用。　　大多數的膠原蛋白都是由二條 (AAB型)或是三條(ABC型)不同胜肽鏈形成的異源三股螺旋結構，因此研究異源三股螺旋結構更能模擬天然的膠原蛋白。我們實驗室先前的研究中，成功的利用cation-π作用力來促使(POG)6(PKG)3與(POG)6(FOG)3胜肽形成AAB型的異源三股螺旋。在第二部分的研究中，我們在 (POG)6(PKG)3的N端先修飾上三個Gly當spacer，再修飾上一個Cys形成CGGG(POG)6(PKG)3，氧化後可以成為雙硫鍵相接的胜肽鏈，此外在CGGG(POG)6(PKG)3上其中一個POG的Gly置換成Sar，因為置換成Sar會使三股螺旋損失氫鍵使結構不穩定，希望藉由雙硫鍵和cation-π作用力形成更穩定的AAB型異源三股螺旋並探討氫鍵對此結構的貢獻。研究結果發現，CGGG(POG)6(PKG)3與(POG)6(PKG)3的性質相似，且與(POG)6(FOG)3混合後會形成異源三股螺旋結構，並觀察到cation-π作用力，但是在氧化後形成雙硫鍵的CGGG(POG)6(PKG)3與CGGG(POG)5 (POSar) (POG) (PKG)3並不能與(POG)6(FOG)3形成類似AAB型的異源三股螺旋結構，顯示位於N端的雙硫鍵並無助於異源三股螺旋的摺疊，而Sar的置換對三股螺旋造成結構很大的破壞。

關鍵字

硼烷共軛化合物；雙硫鍵；異源三股螺旋

並列摘要

Collagen is the most abundant protein in mammals and is an important protein in human body. Collagen has been widely used in biomedical materials because of its biocompatibility, low immunity, and biodegradability. In the first part of this thesis, we prepared the collagen mimetic peptides (POG)7-boroncage conjugate, and investigated the delivery of boron into cells for the potential application in boron neutron capture therapy (BNCT). The results show that the boroncage moiety only slightly destabilize the collagen triple helix and the peptide-boroncage conjugate can enter cells. Most of natural collagens are heterotrimers composed of two (AAB) or three (ABC) different peptide chains, and thus studying heterotrimers can better mimic natural collagens. In our early study, we showed that (POG)6(PKG)3 can form heterotrimers with (POG)6(FOG)3 via cation-π interaction. Here we futher added Cys into the N-terminus of (POG)6(PKG)3, CGGG(POG)6(PKG)3, and replaced Gly to Sar, CGGG(POG)5(POSar)(POG) (PKG)3 to study the effects of disulfides and hydrogen bonding on heterotrimers. The results reveal that CGGG(POG)6(PKG)3 is similar to (POG)6(PKG)3 can form AAB-type heterotrimers with (POG)6(FOG)3, and the cation-π interactions could induce the AAB-type heterotrimers. However, neither disulfide-linked CGGG(POG)6(PKG)3 nor disulfide-linked CGGG(POG)5(POSar) (POG)(PKG)3 can form AAB-type heterotrimers with (POG)6(FOG)3. The resulted showed that the disulfide bond at the N-terminus could not facilitate to the folding of collagen heterotrimers and the replacement of Gly to Sar significantly destabilized the triple helix.

並列關鍵字

borane conjugate ； disulfides ； heterotrimers

參考文獻

1. Cowan, P. M.; McGavin, S.; NORTH, A. C. T., The polypeptide chain configuration of collagen. Nature 1955, 176, 1062-1064.

2. Rich, A.; Crick, F. H. C., The Structure of collagen. Nature 1955, 195, 915-916.

3. Rich, A.; Crick, F. H. C., The molecular structure of collagen. J. Mol. Biol. 1961, 3, 483-484.

4. Bella, J.; Brodsky, B.; Berman, H. M., Hydration structure of a collagen peptide. Structure 1995, 3, 893-906.

5. Hinderaker, M. P.; Raines, R. T., An electronic effect on protein structure. Protein Sci. 2003, 12, 1188-1194.

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膠原蛋白模擬胜肽與硼烷共軛化合物的製備及雙硫鍵與cation-π作用力對膠原蛋白異源三股螺旋摺疊探討

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