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Proteomic Investigation of Murine Neuronal α7-Nicotinic Acetylcholine Receptor Interacting Proteins.
Mulcahy, Matthew J; Paulo, Joao A; Hawrot, Edward.
Afiliação
  • Mulcahy MJ; Division of Biology and Biological Engineering , California Institute of Technology , 1200 East California Boulevard , Pasadena , California 91125-2900 , United States.
  • Paulo JA; Department of Molecular Pharmacology, Physiology and Biotechnology , Brown University , Providence , Rhode Island 02912 , United States.
  • Hawrot E; Department of Cell Biology , Harvard Medical School , 240 Longwood Avenue , Boston , Massachusetts 02115 , United States.
J Proteome Res ; 17(11): 3959-3975, 2018 11 02.
Article em En | MEDLINE | ID: mdl-30285449
The α7-nicotinic acetylcholine receptor (α7-nAChR) is a ligand-gated ion channel that is expressed widely in vertebrates and is the principal high-affinity α-bungarotoxin (α-bgtx) binding protein in the mammalian CNS. α7-nAChRs associate with proteins that can modulate its properties. The α7-nAChR interactome is the summation of proteins interacting or associating with α7-nAChRs in a protein complex. To identify an α7-nAChR interactome in neural tissue, we isolated α-bgtx-affinity protein complexes from wild-type and α7-nAChR knockout (α7 KO) mouse whole brain tissue homogenates using α-bgtx-affinity beads. Affinity precipitated proteins were trypsinized and analyzed with an Orbitrap Fusion mass spectrometer. Proteins isolated with the α7-nAChR specific ligand, α-bgtx, were determined to be α7-nAChR associated proteins. The α7-nAChR subunit and 120 additional proteins were identified. Additionally, 369 proteins were identified as binding to α-bgtx in the absence of α7-nAChR expression, thereby identifying nonspecific proteins for α7-nAChR investigations using α-bgtx enrichment. These results expand on our previous investigations of α7-nAChR interacting proteins using α-bgtx-affinity bead isolation by controlling for differences between α7-nAChR and α-bgtx-specific proteins, developing an improved protein isolation methodology, and incorporating the latest technology in mass spectrometry. The α7-nAChR interactome identified in this study includes proteins associated with the expression, localization, function, or modulation of α7-nAChRs, and it provides a foundation for future studies to elucidate how these interactions contribute to human disease.
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Texto completo: 1 Base de dados: MEDLINE Assunto principal: Encéfalo / Bungarotoxinas / Proteoma / Subunidades Proteicas / Mapeamento de Interação de Proteínas / Receptor Nicotínico de Acetilcolina alfa7 Idioma: En Ano de publicação: 2018 Tipo de documento: Article

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Encéfalo / Bungarotoxinas / Proteoma / Subunidades Proteicas / Mapeamento de Interação de Proteínas / Receptor Nicotínico de Acetilcolina alfa7 Idioma: En Ano de publicação: 2018 Tipo de documento: Article