酵素在醣類的合成，水解及重組

Enzyme in carbohydrate synthesis, hydrolysis and remodeling

蔡宗義(Tsung-I Tsai)

指導教授 ： 翁啟惠

國立臺灣大學/生物資源暨農學院/生物科技研究所/博士(2014年)

https://doi.org/10.6342/NTU.2014.11138

摘要

醣類,脂質,蛋白質為生命組成的三大基本元素.其中,醣類的特性最為複雜.相較於DNA及protein,醣類具有較多的單醣組成,多元的連結可能及複雜的空間構型.此外,醣類還會連結到DNA, protein及脂類,如同本身的被修飾,構成了生物分子的多樣性.而在癌細胞生物學的領域,有幾個重要的醣脂質Gb5, Globo H及SSEA4,已被證明會大量表達在細胞的表面.目前的一些研究報導指出,相較與常見的蛋白質標靶, 這些醣分子更適合開發成為癌症的標靶治療.然而,因為化學合成上的一些限制與困境,導致大量製備上的困難.因此,發展更有效率的合成方法有高度的急迫性.
在本篇論文當中,我集中在兩個部份.一:藉由原位核苷醣再生技術搭配化學酵素法來合成醣分子疫苗Gb5, Globo H, 及SSEA4. 這個方式提供了一個新的方法來大規模合成寡醣,只需三個純化步驟.二:利用新的岩藻醣水解酶來製備無核心岩藻醣的醣蛋白,此外,再搭配其他內切水解酶的情況之下,可用來判斷岩藻醣的位置.之後,利用glycosynthase可以進一步合成無核心岩藻醣的均質化醣蛋白.此舉有助於醣類結構活性的學習.

關鍵字

酵素 ； 醣類合成 ； 醣類水解 ； 醣類重組 ； 醣分子疫苗

並列摘要

Carbohydrate, lipid and protein are basic components of the life. Among these, carbohydrate is significantly complicated than others: more basic units, many possibility of linkages, and complex steric existence. Furthermore, carbohydrate conjugated to DNA, protein and lipid, as well as the modification of carbohydrate itself, also create the biological diversify. Currently, several studies already showed the glycolipid molecules: Gb5, Globo H and SSEA4, are more suitable for target therapy than traditional protein cancer marker. However, due to the difficult and low yield of chemical synthesis, the development of efficient synthetic methodologies for complex glycans preparation has therefore been in high demand.
In this thesis, I focus on two parts. First part is the chemoenzymatic synthesis of carbohydrate vaccine Gb5, Globo H, and SSEA4 by in situ sugar nucleotide regeneration. This method paves a new way to a large scale synthesis of complicated oligosaccharide in just three purification steps, superior to the tedious and laborious traditional chemical methods. The second part is the discovery of new fucosidase for core-fucose eradication and fucose position determination. Subsequently, the core-fucose free glycoprotein can be further utilized for homogeneous glycoprotein preparation by glycosynthase. This way opens a window for gSAR (glycan structure-activity relationship) study.

並列關鍵字

Enzyme ； carbohydrate synthesis ； carbohydrate hydrolysis ； carbohydrate remodeling ； carbohydrate vaccine

參考文獻

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