Antibodies to synthetic peptides as probes for the binding site on the alpha subunit of the acetylcholine receptor.

Neumann, Drorit; Gershoni, Jonathan M.; Fridkin, Mati; Fuchs, Sara

doi:10.1073/pnas.82.10.3490

Cited by 83 publications

(49 citation statements)

References 28 publications

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“…It is clear that the location of the sequence does not guarantee accessibility to an antibody, since interactions with adjacent subunits may shield the sequence which matches the antigenic peptide. Antibodies to the peptides a1 -20 and a125 -143 interacted more with reduced AChR than with AChR, results consistent with an influence of the 192-193 disulfide on the conformation of the receptor [152].…”

Section: Desensitization and Resensitizationsupporting

confidence: 53%

A structural and dynamic model for the nicotinic acetylcholine receptor

Kosower

1987

European Journal of Biochemistry

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Folding of the five polypeptide subunits (azpy6) of the nicotinic acetylcholine receptor (AChR) into a functional structural model is described. The principles used to arrange the sequences into a structure include: (1) hydrophobicity --f membrane-crossing segments; (2) amphipathic character + ion-carrying segments (ion channel with single group rotations); (3) molecular shape (elongated, pentagonal cylinder) + folding dimensions of exobilayer portion; (4) choice of acetylcholine binding sites + specific folding of exobilayer segments; (5) location of reducible disulfides (near agonist binding site) -+ additional specification of exobilayer arrangement; (6) genetic homology + consistency of functional group choices; (7) noncompetitive antagonist labeling + arrangement of bilayer helices. The AChR model is divided into three parts: (a) exobilayer consisting of 11 antiparallel p-strands from each subunit; (b) bilayer consisting of four hydrophobic and one amphiphilic a-helix from each subunit; (c) cytoplasmic consisting of one (folded) loop from each subunit.The exobilayer strands can form a closed 'flower' (the 'resting state') which is opened ('activated') by agonists bound perpendicular to the strands. Rearrangement of the agonists to a strand-parallel position and partial closing of the 'flower' leads to a desensitized receptor. The actions of acetylcholine and succinoyl and suberoyl bis-cholines are clarified by the model. The opening and closing of the exobilayer 'flower' controls access to the ion channel which is composed of the five amphiphilic bilayer helices. A molecular mechanism for ion flow in the channel is given. Openings interrupted by short duration closings (50 ps) depend upon channel group motions.The unusual photolabeling of intrabilayer serines in a, p and 6 subunits but not in y subunits near the binding site for non-competitive antagonists is explained along with a mechanism for the action of these antagonists such as phencyclidine. The unusual a 192Cys-193Cys disulfide may have a special peptide arrangement, such as a cispeptide bond to a following proline (G.A. Petsko and E.M. Kosower, unpublished results). The position of phosphorylatable sites and proline-rich segments are noted for the cytoplasmic loops.The dynamic behavior of the AChR channel and many different experimental results can be interpreted in terms of the model. An example is the lowering of ionic conductivity on substitution of bovine for Torpedo 6 M2 segment. The model represents a useful construct for the design of experiments onAChR. perimenter into a better understanding of current information and to the formulation of new approaches to the problem. In the absence of a definitive structural determination, the structure and dynamic properties of a large molecular complex like the acetylcholine receptor (AChR) must be inferred from a great variety of facts aided by physical and chemical theory. Even if the elusive crystal structure for AChR were obtained, theories for the dynamic behavior would still be necessary. I have...

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Section: Desensitization and Resensitizationsupporting

confidence: 53%

A structural and dynamic model for the nicotinic acetylcholine receptor

Kosower

1987

European Journal of Biochemistry

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“…In the group of HLA-DR5 healthy controls, the distribution was different, with 3 of 14 (21%) responding and 11 of 14 not responding to p195-212 (P < 0.007). Non- (6,(12)(13)(14)(15). In this report, seven peptides that represent sequences of the human AChR a-subunit and one peptide with a region of the Torpedo AChR (Table I) (19,20).…”

Section: Resultsmentioning

confidence: 99%

“…The sequences of the different peptides are shown in Table I. Peptide synthesis and characterization were carried out as previously described (6,7).…”

Section: Methodsmentioning

confidence: 99%

In vitro proliferative responses and antibody titers specific to human acetylcholine receptor synthetic peptides in patients with myasthenia gravis and relation to HLA class II genes.

Brocke¹,

Brautbar²,

Steinman³

et al. 1988

J. Clin. Invest.

Self Cite

120

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To investigate which parts of the acetylcholine receptor are involved in the initiation and development of myasthenia gravis (MG), peptides representing different sequences of the human acetylcholine receptor a-subunit were synthesized. These peptides were tested for their ability to stimulate T cells of myasthenic patients and healthy control patients in proliferation assays and to bind to sera antibodies. Three of eight peptides discriminated significantly between the two groups in the proliferation assay, as well as in their ability to bind to serum antibodies. HLA-DR3 and DR5 were associated with proliferative responses to specific AChR peptides in the group of myasthenics. Acetylcholine receptor epitopes that might play a specific role in myasthenia gravis thus were demonstrated.

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“…The primary structures of the polypeptide chains have been deduced from the precursorcDNA sequences and various structural models of the protein complex in its membrane environment and of the ion channel formed by the receptor protein have been proposed (review [l]). The binding sites for agonists and competitive antagonists are located on the a-subunits, probably around position 192/193 of the primary structure [2,3]. The location and structure of the ion channel are unknown.…”

Section: Introductionmentioning

confidence: 99%

The ion channel of the nicotinic acetylcholine receptor is formed by the homologous helices M II of the receptor subunits

1986

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A binding site for the channel-blocking noncompetitive antagonist [3H]triphenylmethylphosphonium ([3H]TPMP') was localized in the cc-, /I-and b-chains of the nicotinic acetylcholine receptor (AChR) from Torpedo marmorata electric tissue. The photolabel was found in homologous positions of the highly conserved sequence helix II, tl 248,j 254, and 6 262. The site of the photoreaction appears to not be affected by the functional state of the receptor. [3H]TPMP+ was found in position 6 262 independent of whether photolabeling was performed with the receptor in its resting, desensitized or antagonist state. A model of the AChR ion channel is proposed, according to which the channel is formed by the five helices II contributed by the five receptor subunits.

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Antibodies to synthetic peptides as probes for the binding site on the alpha subunit of the acetylcholine receptor.

Cited by 83 publications

References 28 publications

A structural and dynamic model for the nicotinic acetylcholine receptor

A structural and dynamic model for the nicotinic acetylcholine receptor

In vitro proliferative responses and antibody titers specific to human acetylcholine receptor synthetic peptides in patients with myasthenia gravis and relation to HLA class II genes.

The ion channel of the nicotinic acetylcholine receptor is formed by the homologous helices M II of the receptor subunits

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