莱茵衣藻腺苷二磷酸葡萄糖焦磷酸化酶的相互作用蛋白组分析

微藻利用光能和CO₂合成淀粉作为藻细胞的储能物质,是以燃料乙醇为代表的液体生物燃料的可持续原料来源.理解微藻淀粉代谢关键酶的催化机制和调控方式是基因工程操作以提高藻细胞淀粉积累能力的前提. 基于此,以莱茵衣藻淀粉合成代谢中的关键酶腺苷二磷酸葡萄糖焦磷酸化酶(AGPase)催化亚基为探针蛋白,获取该酶的GST融合蛋白后使用GST沉降分析实验结合HPLC-MS/MS的蛋白质检测技术及生物信息学分析方法获得一组与AGPase催化亚基存在相互作用蛋白质,主要包含分子伴侣蛋白、光合作用相关蛋白和信号蛋白分子等. 通过综合分析上述蛋白组的生理功能及亚细胞定位,揭示了它们通过与AGPase相互作用在淀粉合成代谢中的生物学意义. 

Interaction of Proteins with ADP-Glucose Pyrophosphorylase from Chlamydomonas Reinhardtii

Starch, synthesized by microalgae with utilization of light energy and carbon dioxide, is the sustainable feedstock for large scale production of fuel ethanol. In order to enhance accumulation of starch by genetic engineering, it is essential to understand catalytic process and regulation of the key enzymes of starch metabolism in microalgae. For this reason, the rate-limiting enzyme of starch biosynthesis, ADP-glucose pyrophosphorylase (AGPase) in Chlamydomonas reinhardtii, was selected as bait protein for protein-protein interaction (PPI) investigation. With acquisition of AGPase-GST fusion protein, GST-pull down assay was introduced to detect the interaction connected with AGPase. The acquired proteins were further separated and analyzed qualitatively by HPLC-MS/MS. Subsequently, the data was mining by bioinformatics tools. Finally, eleven kinds of proteins were speculated to interact with AGPase, including molecular chaperones, photosynthesis related proteins and signaling molecules. The biological function behind the interaction between these proteins and AGPase can be inferred from the analysis of their subcellular locations and physiological feature.