新穎類似五環醣生合成基因的蛋白質表現與功能性鑑定

Abstract

許多含醣天然物皆具有重要的生物活性，例如抗腫瘤、抗細菌、抗真菌、抗寄生蟲等，且這些天然物通常含有去氧醣的修飾，而醣基的存在往往對於天然物生物活性有重要的影響。因此，利用生合成酵素來改變醣類結構以影響生物活性為當前科學家們的目標之一。本研究探討新穎類似五環醣生成基因，並進行此基因的蛋白質表現與功能性鑑定。 我們在大腸桿菌中進行異體表達，將其表達條件最佳化之後，以親和性管柱純化類似五環醣生合成酵素，並藉由十二烷基硫酸鈉聚丙烯酰胺凝膠電泳分析純化結果。為了針對純化後之類似五環醣生合成酵素進行功能性鑑定，我們以可能的醣類受質進行酵素反應，並分析此類似五環醣生合成酵素對還原輔因子NAD(P)H之特性。本研究探討此類似五環醣生合成基因的功能性分析，以提供科學家更多的資訊來探討醣類生合成的相關基因。

Many natural products with biological activities, including antitumor, antibiotic and antifungal activities, contain sugar moieties. These sugar moieties often play important roles in activity of their parent compounds. The modification or removal of the sugar moieties often changes the reported activities. In this study, we choose a novel gene similar to furanose biosynthetic gene as a research target, which has been speculated to participate in the biosynthetic pathway of furanose or related glycosides. By heterologous expression in Escherichia coli, the expression condition of the protein encoded by the gene was optimized. Then, this enzyme was purified by affinity column and analyzed with SDS-PAGE. In order to characterize the enzymatic function, a spectophotometric method was used to detect enzyme activity. In this study, this enzyme was analyzed with possible cofactors, NADPH and NADH. The results thus provide information about this novel enzyme possibly participating in biosynthetic pathways of furanose or related glycosides.