- 期刊
Elimination of False Negative Results in the Two-Hybrid System in the Phagocyte NADPH Oxidase
利用酵母菌Two-hybrid 系統研究吞噬細胞的NADPH Oxidase的組成分子及其偽陰性反應
摘要
酵母菌Two-hybrid系統近年來已被廣泛做用於偵測蛋白質與蛋白質之間的連繫。此方法之基本原理係將兩種不同的polypeptides分別融合（fuse）到一個transcrptional activator的DNA-binding domain和activation domain上，藉由報告基因（reporters）之表達以偵測兩種蛋白質之間的關連與否。本文是首次成功地利用Two-hybrid系統來研究吞噬細胞的NADPH Oxidase在胞內的兩個組成分子，並探討此方法在使用之方向性（orientation）及偽陰性（false negative）。
關鍵字
無資料並列摘要
The yeast two-hybrid system is finding increased use in the study of interactions between proteins. In this method, two polypeptides are expressed in yeast as fusion proteins to a transcriptional activator DNA-binding domain (bd) and activating domain (ad), respectively. Interaction between the two polypeptides reconstitutes function of a transactivator which controls expression of reporters. The phagocyte NADPH oxidase is a complex of membrane cytochrome b558 (comprised of subunits p22-phox and gp9l-phox) and three cytosol proteins (p47-phox, p67-phox, and p2lrac) that translocate to membrane and bind to cytochrome b558. This is the first report to demonstrate that two of cytosolic components of cytochrome b558, p47-phox binding to p67-phox each other. We encountered several methodological problems in the two-hybrid system which are the focus of this report.