Proteomic analysis of an alpha7 nicotinic acetylcholine receptor interactome.
The alpha7 nicotinic acetylcholine receptor (nAChR) is well established as the principal high-affinity alpha-bungarotoxin-binding protein in the mammalian brain. We isolated carbachol-sensitive alpha-bungarotoxin-binding complexes from total mouse brain tissue by affinity immobilization followed by selective elution, and these proteins were fractionated by SDS-PAGE. The proteins in subdivided gel lane segments were tryptically digested, and the resulting peptides were analyzed by standard mass spectrometry. We identified 55 proteins in wild-type samples that were not present in comparable brain samples from alpha7 nAChR knockout mice that had been processed in a parallel fashion. Many of these 55 proteins are novel proteomic candidates for interaction partners of the alpha7 nAChR, and many are associated with multiple signaling pathways that may be implicated in alpha7 function in the central nervous system. The newly identified potential protein interactions, together with the general methodology that we introduce for alpha-bungarotoxin-binding protein complexes, form a new platform for many interesting follow-up studies aimed at elucidating the physiological role of neuronal alpha7 nAChRs.
Paulo
Joao A
JA
Graduate Program in Molecular Biology, Cell Biology and Biochemistry and Department of Molecular Pharmacology, Physiology and Biotechnology, Brown University, Providence, Rhode Island 02912, USA.
Brucker
William J
WJ
Hawrot
Edward
E
D000818:Animals; D001921:Brain / Q000378:metabolism*; D002038:Bungarotoxins / Q000378:metabolism*; D013058:Mass Spectrometry; D051379:Mice; D018345:Mice, Knockout; D011485:Protein Binding; D020543:Proteome / Q000378:metabolism*; D011978:Receptors, Nicotinic / Q000378:metabolism*; D064569:alpha7 Nicotinic Acetylcholine Receptor
D002038:Bungarotoxins; C579166:Chrna7 protein, mouse; D020543:Proteome; D011978:Receptors, Nicotinic; D064569:alpha7 Nicotinic Acetylcholine Receptor