通过光敏保护策略半合成糖蛋白肿瘤坏死因子α

通过表达蛋白质连接（expressed protein ligation，EPL）对蛋白质进行半合成的策略相较于全合成有着效率高、易操作的优势。然而，由于半胱氨酸脱硫具有无选择性的特性，自然化学连接（NCL）/脱硫方法学组无法被应用于构建有多个半胱氨酸（Cys）位点的蛋白质EPL半合成，限制了EPL在蛋白质合成领域中的应用。本项目通过蛋白质后期修饰的概念在表达蛋白质上引入半胱氨酸的光敏保护基，对蛋白质进行正交保护/NCL/脱硫/光引发脱保护，对含有多个Cys的蛋白质通过丙氨酸位点进行半合成，进而拓展自然化学连接方法学组在EPL半合成领域中的应用。该技术将被应用于合成具有单一糖型的肿瘤坏死因子α蛋白质家族，并通过细胞实验和肿瘤小鼠实验研究寡糖对该细胞因子生物活性的影响。

In comparison of protein total synthesis, semi-synthesis via expressed protein ligation (EPL) is more efficient and easy-operating. However, due to lack of selectivity of desulfurization of cysteine residues, native chemical ligation (NCL)/desulfurization methodology cannot be applied to construct proteins bearing multiple cysteines, which limits the potential application of EPL in protein synthesis field. Through introducing a photo-labile protecting group to expressed protein, which adapted the concept of late-stage modification, this project aims to realize EPL ligation at alanine junction via orthogonal-protecting/EPL/desulfurization/photo-driven deprotection protocol, and therefore expands the potential application of NCL methodologies in protein semi-synthesis field. In addition, this novel technology will be applied to the semi-synthesis of homogeneously glycosylated Tumour Necrosis Factor α. The synthetic protein standards will be used as probes for cell studies and tumour mice model experiments to reveal the influence caused by oligosaccharides to its cytokine activities.