Comparison of the subunit structure of acetylcholine receptors from muscle and electric organ of Electrophorus electricus

Gullick, William J.; Lindstrom, Jon T.

doi:10.1021/bi00285a014

Cited by 16 publications

(2 citation statements)

References 44 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Mouse monoclonal anti-␣-AChR subunit was purchased from Transduction Laboratories (BD Biosciences). Antibody to the ␤-AChR subunit (mAb124) was a gift from J. Lindstrom, University of Pennsylvania, Philadelphia, PA (Gullick and Lindstrom, 1983). Rabbit anti-pAChR␤1 (Y390) antiserum (sc-17087R) was obtained from Santa Cruz Biotechnology.…”

Section: Methodsmentioning

confidence: 99%

ColQ Controls Postsynaptic Differentiation at the Neuromuscular Junction

Sigoillot¹,

Bourgeois²,

Lambergeon³

et al. 2010

J. Neurosci.

View full text Add to dashboard Cite

Section: Methodsmentioning

confidence: 99%

ColQ Controls Postsynaptic Differentiation at the Neuromuscular Junction

Sigoillot¹,

Bourgeois²,

Lambergeon³

et al. 2010

J. Neurosci.

View full text Add to dashboard Cite

“…Further dissection of the receptor can be achieved through the preparation of peptide fragments by limited proteolysis of isolated subunits. This approach has been employed, for example, to compare the subunit structures of AcChoR from different sources (11,12) and to characterize the domain specificity of anti-AcChoR monoclonal antibodies (13). One difficulty in utilizing parts of the AcChoR to study receptor function is that as the AcChoR is dismantled its characteristic functions may become less recognizable or no longer detectable.…”

mentioning

confidence: 99%

Binding of alpha-bungarotoxin to proteolytic fragments of the alpha subunit of Torpedo acetylcholine receptor analyzed by protein transfer on positively charged membrane filters.

Wilson¹,

Gershoni²,

Hawrot³

et al. 1984

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Proteolytic fragments of the a subunit of the acetylcholine receptor retain the ability to bind a-bungarotoxin following resolution by polyacrylamide gel electrophoresis and immobilization on protein transfers. The a subunit of the acetylcholine receptor of Torpedo electric organ was digested with four proteases: Staphylococcus aureus V-8 protease, papain, bromelain, and proteinase K. The proteolytic fragments resolved on 15% polyacrylamide gels were electrophoretically transferred onto positively charged nylon membrane filters. When incubated with 0.3 nM 1251-labeled a-bungarotoxin and autoradiographed, the transfers yielded patterns of labeled bands characteristic for each protease. The molecular masses of the fragments binding toxin ranged from 7 to 34 kDa, with major groupings in the 8-, 18-, and 28-kDa ranges. The apparent affinity of the fragments for a-bungarotoxin as determined from the IC50 value was 6.7 X 10-8 M. The labeling of fragments with a-bungarotoxin could be inhibited by prior affinity alkylation of receptor-containing membranes with 4-(N-maleimido)-a-benzyltrimethylammonium iodide. These findings demonstrate that immobilized proteolytic fragments as small as 1/5 the size of the a subunit retain the structural characteristics necessary for binding a-bungarotoxin, although the toxin is bound to the fragments with lower affinity than to the native receptor. The effect of affinity ligand alkylation demonstrates that the a-bungarotoxin binding site detected on the proteolytic fragments is the same as the affinitylabeled acetylcholine binding site on the intact acetylcholine receptor.The nicotinic acetylcholine receptor (AcChoR) that functions to transduce a chemical signal into an electrical event in many neuronal systems has been well characterized both physiologically and biochemically (for review, see refs. 1 and 2). The AcChoR purified from the electroplaque of the electric ray Torpedo californica consists of four subunits present in the molar stoichiometry of a2/3y8 (3)(4)(5). This pentameric complex contains the agonist binding sites responsible for receptor activation and desensitization as well as the ion channel for cation flux through the membrane. Recent information on the primary amino acid sequences of the polypeptides comprising the receptor (5-9) provides a basis for analyzing and understanding these receptor functions at the molecular level.One approach in elucidating receptor function is the dismantling of the intact receptor into its constituent parts and determination of the functions and properties of these simpler components. For example, the four subunits of the AcChoR have been isolated by gel electrophoresis in NaDodSO4 and tested for a-bungarotoxin (a-BTX) binding after

show abstract

Brain and Muscle Nicotinic Receptors: Complex Homologous Proteins Carrying Multiple Binding Sites

Conti‐Tronconi

Raftery

1987

Tobacco Smoking and Nicotine

View full text Add to dashboard Cite

Comparison of the subunit structure of acetylcholine receptors from muscle and electric organ of Electrophorus electricus

Cited by 16 publications

References 44 publications

ColQ Controls Postsynaptic Differentiation at the Neuromuscular Junction

ColQ Controls Postsynaptic Differentiation at the Neuromuscular Junction

Binding of alpha-bungarotoxin to proteolytic fragments of the alpha subunit of Torpedo acetylcholine receptor analyzed by protein transfer on positively charged membrane filters.

Brain and Muscle Nicotinic Receptors: Complex Homologous Proteins Carrying Multiple Binding Sites

Contact Info

Product

Resources

About