Biomaterial Database

Photoaffinity labeling of the acetylcholine transporter. 1992

G A Rogers, and S M Parsons

Department of Chemistry, University of California, Santa Barbara 93106.

The acetylcholine (AcCh) binding site in the AcCh transporter-vesamicol receptor (AcChT-VR) present in synaptic vesicles isolated from the electric organ of Torpedo was characterized. A high-affinity analogue of AcCh containing an aryl azido group, namely, cyclohexylmethyl cis-N-(4-azidophenacyl)-N-methylisonipecotate bromide (AzidoAcCh), was synthesized in nonradioactive and highly tritiated forms. AzidoAcCh was shown to be a competitive inhibitor of [3H]AcCh active transport and binding of [3H]-vesamicol to the allosteric site. The [3H]AzidoAcCh saturation curve was determined. In all cases the AcChT.AzidoAcCh complex exhibited an inhibition or dissociation constant of about 0.3 microM. Binding of [3H]AzidoAcCh was inhibited by vesamicol and AcCh. AzidoAcCh irreversibly blocked greater than 90% of the [3H]vesamicol binding sites after multiple rounds of photolysis and reequilibration with fresh ligand. Autofluorographs of synaptic vesicles photoaffinity-labeled with [3H]AzidoAcCh showed specific labeling of material exhibiting a continuous distribution from 50 to 250 kDa after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The result demonstrates that the AcChT has an unexpected structure highly suggestive of the synaptic vesicle proteoglycan.

UI	MeSH Term	Description	Entries
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D010777	Photochemistry	A branch of physical chemistry which studies chemical reactions, isomerization and physical behavior that may occur under the influence of visible and/or ultraviolet light.	Photochemistries
D010880	Piperidines	A family of hexahydropyridines.
D002352	Carrier Proteins	Proteins that bind or transport specific substances in the blood, within the cell, or across cell membranes.	Binding Proteins,Carrier Protein,Transport Protein,Transport Proteins,Binding Protein,Protein, Carrier,Proteins, Carrier
D004557	Electric Organ	In about 250 species of electric fishes, modified muscle fibers forming disklike multinucleate plates arranged in stacks like batteries in series and embedded in a gelatinous matrix. A large torpedo ray may have half a million plates. Muscles in different parts of the body may be modified, i.e., the trunk and tail in the electric eel, the hyobranchial apparatus in the electric ray, and extrinsic eye muscles in the stargazers. Powerful electric organs emit pulses in brief bursts several times a second. They serve to stun prey and ward off predators. A large torpedo ray can produce of shock of more than 200 volts, capable of stunning a human. (Storer et al., General Zoology, 6th ed, p672)	Electric Organs,Organ, Electric,Organs, Electric
D004591	Electrophoresis, Polyacrylamide Gel	Electrophoresis in which a polyacrylamide gel is used as the diffusion medium.	Polyacrylamide Gel Electrophoresis,SDS-PAGE,Sodium Dodecyl Sulfate-PAGE,Gel Electrophoresis, Polyacrylamide,SDS PAGE,Sodium Dodecyl Sulfate PAGE,Sodium Dodecyl Sulfate-PAGEs
D000109	Acetylcholine	A neurotransmitter found at neuromuscular junctions, autonomic ganglia, parasympathetic effector junctions, a subset of sympathetic effector junctions, and at many sites in the central nervous system.	2-(Acetyloxy)-N,N,N-trimethylethanaminium,Acetilcolina Cusi,Acetylcholine Bromide,Acetylcholine Chloride,Acetylcholine Fluoride,Acetylcholine Hydroxide,Acetylcholine Iodide,Acetylcholine L-Tartrate,Acetylcholine Perchlorate,Acetylcholine Picrate,Acetylcholine Picrate (1:1),Acetylcholine Sulfate (1:1),Bromoacetylcholine,Chloroacetylcholine,Miochol,Acetylcholine L Tartrate,Bromide, Acetylcholine,Cusi, Acetilcolina,Fluoride, Acetylcholine,Hydroxide, Acetylcholine,Iodide, Acetylcholine,L-Tartrate, Acetylcholine,Perchlorate, Acetylcholine
D000345	Affinity Labels	Analogs of those substrates or compounds which bind naturally at the active sites of proteins, enzymes, antibodies, steroids, or physiological receptors. These analogs form a stable covalent bond at the binding site, thereby acting as inhibitors of the proteins or steroids.	Affinity Labeling Reagents,Labeling Reagents, Affinity,Labels, Affinity,Reagents, Affinity Labeling
D000818	Animals	Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA.	Animal,Metazoa,Animalia
D001386	Azides	Organic or inorganic compounds that contain the -N3 group.	Azide