Analysis of membrane protein clusters using fluorescence lifetime FRET

为了使用荧光寿命FRET监测膜蛋白的聚类，在细胞中表达GFP或RFP标记的Tat 2，其编码酵母色氨酸通透酶Tat 2。然而，荧光信号太低，无法获得有效的光子计数。修改策略，进行非变性PAGE来阐明Tat 2簇。大多数的Tat 2蛋白质似乎迁移后，膜馏分适度处理的非离子型洗涤剂作为单体。利用高压时间分辨荧光各向异性测量系统，分析了深海耐压生物质膜的流动性。膜显示是刚性的，而不是一般的概念，深海微生物的膜是高度流动的。

To monitor clustering of the membrane protein using fluorescence lifetime FRET, GFP-or RFP-tagged Tat2, which encodes yeast tryptophan permease Tat2, was expressed in cells. However, the fluorescence signal was too lowto obtain operative photon counting. Revisingthe strategy, native PAGE was performed toelucidateTat2 clusters. The majority of Tat2 proteins appeared to migrate as monomers after the membrane fraction was moderately treated with a non-ionic detergent. By means of high-pressure time-resolved fluorescence anisotropy measurement system, fluidity of the plasma membrane was analyzed in a deep-sea piezophile. The membrane revealed to be rigid as opposed to a general concept that the membrane of deep-seamicrobes is highly fluid.