PPIs Detection Reagent : Fluoppi [ p53-MDM4 ]

Fluoppi is a technology providing an easy way to visualize protein-protein interactions (PPIs) with a high signal to noise ratio. It employs an oligomeric assembly helper tag (Ash-tag) and a tetrameric fluorescent protein tag (FP-tag) to create detectable fluorescent puncta when there are interactions between two proteins fused to the tags. Schematic images are illustrated in Figure 1, where genetic fusion of protein X with FP-tag, and Y with Ash-tag creates a tetrameric fluorescent fusion protein X-FP and an oligomeric fusion protein Y-Ash respectively. Because each fusion protein has multiple Xs or Ys, the interaction between X and Y causes phase-separated droplets where the fluorescence by X-FP is concentrated and detectable as fluorescent puncta.

This product contains two expression cassettes for detecting p53-MDM4 interaction in living cells. One encodes a fusion protein Ash/p53, and the other encodes humanized Azami-Green (hAG)/MDM4. MDM4 is also known as MDMX. Partial sequences responsible for this interaction are used for this product. Co-transfection of DNA cassettes, Ash/p53 and hAG/MDM4, results in formation of cytoplasmic fluorescent puncta. After addition of p53-MDM4 PPI inhibitors, the puncta disappeared within a day, indicating the p53-MDM4 complex was disrupted.