合成硫酸化之乙醯乳醣胺四醣分子： 瞭解硫酸根基團如何影響半乳醣凝集素的結合力

Abstract

半乳醣凝集素 (galectin) 是一種能和醣體有良好作用力的醣接合蛋白，其醣辨識區 (carbohydrate recognition domain, CRD) 是由相似的氨基酸序列所組成，且能夠辨識具有β-半乳糖苷 (β-galactoside) 的醣體。近期的研究也證實半乳醣凝集素在許多疾病和生物活性上扮演相當重要的角色，如半乳糖凝集素-1可以導致漿細胞不正當分化、促使腸病毒71型的感染和促進口腔癌細胞的轉移等。 本實驗室先前針對具有硫酸化的N-乙醯乳醣胺及其衍生物進行結合能力試驗發現，在N-乙醯乳醣胺的Gal的3號羥基及GlcNAc的6號羥基修飾上硫酸根基團，會對半乳糖凝集素的結合能力有明顯提升。且由文獻可得知，以第二型N-乙醯乳醣胺以β-1,3醣苷鍵為重複單元所組成長鏈寡醣分子對於半乳糖凝集素-1的親合能力有顯著的提升。 因此，本論文的主題為合成Gal-β1,4-GlcNAc-β1,3-Gal-β-1,4-GlcNAc的四醣骨架，並在其還原端 Gal的3號羥基位置修飾硫酸根基團及兩個GlcNAc的6號羥基位置修飾上硫酸根基團，進而去探討電荷及鏈長對於半乳糖結合能力的影響。合成此類化合物將涉及有機醣化學合成中的 (1) 醣予體(glycosyl acceptor)/醣受體 (glycosyl donor) 單元的設計與合成； (2) 醣苷鍵 (glycosidic bond) 的建立；(3) 最佳化保護/去保護的條件； (4) 選擇性在醣類一級醇或二級醇的硫酸化。在合成策略上，設計出具有交錯保護基特性的醣予體及醣受體，運用鄰近基效應 (neighboring group participation) 在醣予體2號碳位置修飾上酯基以建立β-醣苷鍵，最後將特定基團去保護再修飾上硫酸根基團。配合各種光譜證據，以上合成已有了初步的成果。

Galectins are a family of β-galactoside-binding proteins to share a high sequence similarity in their carbohydrate recognition domains. They play an important role in numerous biological activities and have been attributed to several diseases. For example, galectin-1 promotes the formation of Ab-secreting plasma cells, interactes with enterovirus 71 during the infection process, and enhances tumor metastasis. In our previously work, several sulfated disaccharides were found to display selective binding with galectins-1 and -8. To study how the negative charges and chain length contribute to the affinity and selectivity, my thesis lays a special emphasis the synthesis of sulfate and Gal-β1,4-GlcNAc (type II) containing tetrasaccharides. We elaborated the use of orthogonal protecting groups and the preparation of the Gal-β1,4-GlcNAc building block with minimal purification steps. We successfully improved and optimized several key reactions, such as the glycosylation, deprotection of trichloroacetyl amide and removal of benzyl ether. Once being available, the synthesized tetrasaccharide products will be measured and quantitated for their binding effinity by using fluorescence polarization (FP), in comparison with the reported standards (i.e. fluorescein-containing LacNAc)