Related Experiment Video
Updated: Feb 15, 2026
10:09
Detection of Detergent-sensitive Interactions Between Membrane Proteins
Published on: March 7, 2018
6.4K
Detection of Membrane Protein Interactions by Cell-based Tango Assays
Yan Yan1,2,3, Ting-Hai Xu1,2,3, Kaleeckal G Harikumar4
1Key Laboratory of Receptor Research, VARI-SIMM Center, Center for Structure and Function of Drug Targets, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
Bio-Protocol
|February 3, 2018
Summary
This study adapted the Tango assay to detect intramembrane protein interactions. Researchers analyzed amyloid precursor protein C99 interactions within live cells using this modified protein-protein interaction assay.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- The Tango assay is a versatile tool for detecting protein-protein interactions.
- It relies on TEV protease cleavage to release a reporter protein into the nucleus.
- Previous applications focused on soluble or membrane-bound interactions.
Purpose of the Study:
- To adapt and validate the Tango assay for analyzing intramembrane protein interactions.
- To investigate the self-interaction of the amyloid precursor protein (APP) C99 region.
- To establish a method for studying protein associations within cellular membranes.
Main Methods:
- Modification of the Tango assay by fusing the APP C99 region to assay components.
- Expression of C99-TEV site-rTA and C99-TEV constructs in live cells.
- Utilizing luciferase reporter gene activity to quantify C99-C99 interactions.
Main Results:
- Demonstrated successful adaptation of the Tango assay for intramembrane interaction studies.
- Provided evidence for C99-C99 interactions within the cell membrane.
- Quantified the interaction strength based on reporter gene activation.
Conclusions:
- The modified Tango assay is effective for studying intramembrane protein-protein interactions.
- The APP C99 region engages in self-interaction within the membrane.
- This assay provides a novel approach to investigate membrane protein associations.
More Related Videos
Related Concept Videos
01:16Introduction to Membrane Proteins
81.7K
The cell membrane, or plasma membrane, is an ever-changing landscape. It is described as a fluid mosaic where various macromolecules are embedded in the phospholipid bilayer. Among the macromolecules are proteins. The protein content varies across cell types. For example, mitochondrial inner membranes contain ~76% protein content, while myelin contains ~18% protein content. Individual cells contain many types of membrane proteins—red blood cells contain over 50—and different cell...
81.7K
01:30Membrane Proteins
30.6K
Plasma membranes have integral transmembrane proteins involved in facilitated transport. These proteins are collectively referred to as transport proteins, and they function as either channels for the material or as carriers themselves. Channel proteins have hydrophilic domains exposed to the intracellular and extracellular fluids and a hydrophilic channel through their core that provides a hydrated opening for solutes to pass through the membrane layers. Passage through the channel allows...
30.6K
01:24Protein Diffusion in the Membrane
5.7K
Proteins show rotational as well as lateral diffusion across the membrane. The lateral diffusion of proteins was confirmed through the cell fusion experiment where mouse and human cells were fused, resulting in hybrid cells. When the human and mouse cells fused, the specific membrane proteins on human and mouse cells were marked with the red and green-fluorescent markers, respectively. Initially, the red and green fluorescence was located on the respective hemisphere of the cell. As time...
5.7K
01:18Detergent Purification of Membrane Proteins
6.5K
Detergents are used to purify the integral proteins of the membrane. The hydrophobic portion of the detergent can replace membrane phospholipids while solubilizing the membrane proteins. When detergent monomers reach a specific concentration in a solution called critical micelle concentration (CMC), they form micelles. Above CMC, the concentration of the detergent monomers remains in equilibrium with the micelle. The number of detergent monomers present in the CMC varies for each detergent, and...
6.5K
01:29GPI Anchoring of Proteins in the ER Membrane
5.6K
GPI-anchoring is a post-translational, reversible protein modification that is ubiquitous in eukaryotes. Such proteins are primarily present on the exoplasmic leaflet of the plasma membrane.
GPI-anchor structure
A sequence of 11 enzymatic reactions results in the synthesis of the complete GPI anchor consisting of a hydrophobic and a hydrophilic portion. The hydrophobic portion comprises phosphatidylinositol, while the hydrophilic part comprises polar groups like phosphoethanolamine,...
GPI-anchor structure
A sequence of 11 enzymatic reactions results in the synthesis of the complete GPI anchor consisting of a hydrophobic and a hydrophilic portion. The hydrophobic portion comprises phosphatidylinositol, while the hydrophilic part comprises polar groups like phosphoethanolamine,...
5.6K
01:45Tail-anchoring of Proteins in the ER Membrane
3.9K
Tail-anchored, or TA, proteins are estimated to make up to 3-5% of membrane proteins found in the eukaryotic cell. Such proteins have a single transmembrane domain located approximately 30 amino acid residues upstream from the C-terminal end. As a result, the signal recognition particle (SRP) cannot guide a TA protein to the ER membrane for cotranslational insertion. Hence, they are integrated into the ER membrane post-translationally using their C-terminal end as the anchor. TA proteins...
3.9K