title: Proteolytic fragments of the nicotinic acetylcholine receptor identified by mass spectrometry: implications for receptor topography.
abstract: A triple-state quadrupole or a tandem quadrupole Fourier-transform mass spectrometer was used to detect and sequence the peptides released by proteolytic cleavage of the acetylcholine receptor (AcChR) from Torpedo californica electroplax. Fragments in mass range up to 3479 daltons were characterized on the above instrumentation and used to determine proteolytically accessible sites on the receptor. These data were consistent with the cleavage points determined for membrane-bound fragments of the same AcChR samples using gas-phase microsequencing. Each subunit of the receptor is readily cleaved near the C-terminus in the region between the proposed transmembrane hydrophobic alpha-helices MIII and MIV. This region includes the putative regulatory phosphorylation sites and the amphipathic alpha-helix. Cleavage is also observed in the N-terminal domain, but occurs much more slowly than in the C-terminal region. No cleavage was detected in the middle third of the receptor, which includes the proposed transmembrane alpha-helices MI and MII. An evaluation of these data in terms of the transmembrane topography of the AcChR peptides is consistent with a synaptic or extracellular disposition for the region between MIII and MIV.
authors.affiliation: Department of Chemistry, University of Virginia, Charlottesville 22901.
authors.forename: C R
authors.initials: CR
authors.lastname: Moore
authors.forename: J R
authors.initials: JR
authors.lastname: Yates
authors.forename: P R
authors.initials: PR
authors.lastname: Griffin
authors.forename: J
authors.initials: J
authors.lastname: Shabanowitz
authors.forename: P A
authors.initials: PA
authors.lastname: Martino
authors.forename: D F
authors.initials: DF
authors.lastname: Hunt
authors.forename: D S
authors.initials: DS
authors.lastname: Cafiso
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000032:analysis; D002846:Chromatography, Affinity; D013058:Mass Spectrometry; D008954:Models, Biological; D008969:Molecular Sequence Data; D011950:Receptors, Cholinergic; D012697:Serine Endopeptidases; D014101:Torpedo; D014357:Trypsin
chemical_list: D011950:Receptors, Cholinergic; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D014357:Trypsin