Related Experiment Video
Updated: Jun 24, 2026
12:48
Purification of the Membrane Compartment for Endoplasmic Reticulum-associated Degradation of Exogenous Antigens in Cross-presentation
Published on: August 21, 2017
Purification of the vesamicol receptor
1Department of Chemistry, University of California, Santa Barbara 93106.
Biochemistry
|June 30, 1992
Summary
Researchers purified the vesamicol receptor (VR), crucial for acetylcholine transport in cholinergic vesicles. The VR copurified with synaptic vesicle proteoglycan (SV1), suggesting a strong association and confirming its role in acetylcholine transport.
Area of Science:
- Neuroscience
- Biochemistry
- Molecular Biology
Background:
- Cholinergic synaptic vesicles contain the vesamicol receptor (VR), essential for acetylcholine (ACh) transport.
- The VR's precise molecular nature and association with vesicle components remained unclear.
Purpose of the Study:
- To purify and characterize the vesamicol receptor (VR) from Torpedo electric organ cholinergic synaptic vesicles.
- To investigate the VR's association with synaptic vesicle proteins and proteoglycans.
Main Methods:
- Solubilization of VR using cholate detergent and stabilization with glycerol/phospholipids.
- Multi-step purification including hydroxylapatite, wheat germ lectin affinity, DEAE anion-exchange, and size exclusion chromatography.
- Biochemical assays including [3H]vesamicol binding, SDS-PAGE, and epitope mapping (SV1, SV2).
Main Results:
- Purified VR with a specific binding of 4400 pmol/mg protein, exhibiting variable aggregation states (210-3500 kDa).
- VR showed heterogeneous electrophoretic mobility (~240 kDa) without distinct polypeptide bands.
- VR copurified with the SV1 epitope (cholinergic synaptic vesicle proteoglycan) and the SV2 epitope.
Conclusions:
- The VR is strongly associated with cholinergic synaptic vesicle proteoglycan.
- Purified VR retained enantioselectivity and ACh-binding capacity, supporting the allosteric model of the ACh transporter.
- A distinct proteoglycan carrying SV1 and SV2 epitopes, independent of VR, was also identified.
Related Concept Videos
01:18Detergent Purification of Membrane Proteins
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...
01:15Receptor Downregulation in MVBs
Multivesicular bodies (MVBs) are mature endosomes that sort ubiquitinated proteins and then fuse with lysosomes to degrade the sorted proteins. Epidermal growth factor (EGF) and its receptor (EGFR) form a complex that can be internalized through endocytosis, sorted into an MVB, and later degraded.
The EGFR can initiate signaling pathways that lead to cell proliferation, migration, and differentiation. Overexpression of EGFR stimulates cells to proliferate. Excessive EGFR activation may...
The EGFR can initiate signaling pathways that lead to cell proliferation, migration, and differentiation. Overexpression of EGFR stimulates cells to proliferate. Excessive EGFR activation may...
01:15Drug Elimination by Renal Route: Tubular Secretion
Once the process of glomerular filtration is completed, blood carrying unfiltered drug molecules traverses through efferent arterioles and makes its way into the peritubular capillaries in the proximal tubule. A variety of carriers play a pivotal role in actively secreting drugs from these peritubular capillaries into the tubular fluid. The organic anion transporter transfers acidic drugs, against an electrochemical gradient, from the peritubular capillaries into the renal tubule cells and...