Related Experiment Video
Updated: Jan 24, 2026
10:36
Purification of Native Complexes for Structural Study Using a Tandem Affinity Tag Method
Published on: July 27, 2016
10.0K
Purification of the Rhodopsin-Transducin Complex for Structural Studies
Yang Gao1, Jon W Erickson2, Richard A Cerione2
1Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY, USA.
Methods in Molecular Biology (Clifton, N.J.)
|June 2, 2019
Summary
We developed a method to purify the visual signaling complex, rhodopsin-transducin, from native membranes. This method yields a stable, pure complex suitable for structural studies and applicable to other G protein-coupled receptors.
Area of Science:
- Biochemistry
- Structural Biology
- Cell Signaling
Background:
- G protein-coupled receptors (GPCRs) are crucial transmembrane proteins and drug targets.
- Structural studies of GPCR-G protein complexes are vital for understanding cellular signaling.
- Visual phototransduction involves the rhodopsin-transducin complex.
Purpose of the Study:
- To present a method for large-scale purification of the native rhodopsin-transducin complex.
- To enable structural and biophysical studies of GPCR-G protein interactions.
- To establish a versatile method applicable to other GPCRs.
Main Methods:
- Purification of rhodopsin-transducin complex directly from native bovine retinal membranes.
- Utilizing native, lipid-modified proteins (palmitoylated rhodopsin, myristoylated/farnesylated transducin subunits).
- Formation of the complex on purified native cell membranes.
Main Results:
- Achieved large-scale purification of a highly pure and stable rhodopsin-transducin complex.
- Demonstrated the importance of specific lipid modifications for complex formation.
- The purified complex is suitable for biophysical and structural analyses.
Conclusions:
- The described method provides a robust approach for purifying native GPCR-G protein complexes.
- This technique facilitates structural studies of visual phototransduction.
- The method is adaptable for other GPCRs expressed in various cell systems.
Related Concept Videos
01:11Channel Rhodopsins
3.1K
Most organisms use photoreceptors to sense and respond to light. Examples of photoreceptors include bacteriorhodopsins and bacteriophytochromes in some bacteria, phytochromes in plants, and rhodopsins in the photoreceptor cells of the vertebral retina. The light-sensitive property of these receptors is because of the bound chromophores, such as bilin in the phytochromes and retinal in the rhodopsins.
Rhodopsins belong to the family of cell surface proteins called G-protein coupled receptors,...
Rhodopsins belong to the family of cell surface proteins called G-protein coupled receptors,...
3.1K
01:32Assembly of Complex Microtubule Structures
2.5K
Complex microtubule structures are present in resting cells and in dividing cells. In resting cells, they are responsible for maintaining the cellular architecture, tracks for intracellular transport, positioning of organelles, assembly of cilia and flagella. They mediate the bipolar spindle assembly for chromosomal segregation and positioning of the cell division plate in dividing cells. The formation of microtubule complex structures depends on the cell type, cell stage, and cell function.
2.5K
02:34Structural Isomerism
21.5K
Isomerism in Complexes
Isomers are different chemical species that have the same chemical formula. Structural isomerism of coordination compounds can be divided into two subcategories, the linkage isomers and coordination-sphere isomers.
Linkage isomers occur when the coordination compound contains a ligand that can bind to the transition metal center through two different atoms. For example, the CN− ligand can bind through the carbon atom or through the nitrogen atom. Similarly, SCN− can...
Isomers are different chemical species that have the same chemical formula. Structural isomerism of coordination compounds can be divided into two subcategories, the linkage isomers and coordination-sphere isomers.
Linkage isomers occur when the coordination compound contains a ligand that can bind to the transition metal center through two different atoms. For example, the CN− ligand can bind through the carbon atom or through the nitrogen atom. Similarly, SCN− can...
21.5K
02:10Additional Subnuclear Structures
5.3K
The eukaryotic nucleus is a double membrane-bound organelle that contains nearly all of the cell’s genetic material in the form of chromosomes. It is rightly called the “brain” of the cell as it shoulders the responsibility of responding to various physiological processes, stress, altered metabolic conditions, and other cellular signals.
The nucleus contains many membrane-less subnuclear organelles or nuclear bodies, such as nucleoli, Cajal bodies, speckles,...
The nucleus contains many membrane-less subnuclear organelles or nuclear bodies, such as nucleoli, Cajal bodies, speckles,...
5.3K
02:41Protein Complex Assembly
16.7K
Proteins can form homomeric complexes with another unit of the same protein or heteromeric complexes with different types. Most protein complexes self-assemble spontaneously via ordered pathways, while some proteins need assembly factors that guide their proper assembly. Despite the crowded intracellular environment, proteins usually interact with their correct partners and form functional complexes.
Many viruses self-assemble into a fully functional unit using the infected host cell to...
Many viruses self-assemble into a fully functional unit using the infected host cell to...
16.7K
01:57Protein Complexes with Interchangeable Parts
2.9K
Groups of proteins may form a complex where each protein in this complex has a different role in the overall execution of the complex’s function. Often some of the proteins in the complex can be replaced by a closely related variant to give a complex that contains many of the same components yet is functionally distinct.
The SCF ubiquitin ligase is a protein complex of five individual proteins. This complex attaches ubiquitin to other target proteins to mark them for degradation. In order...
The SCF ubiquitin ligase is a protein complex of five individual proteins. This complex attaches ubiquitin to other target proteins to mark them for degradation. In order...
2.9K