Topological dispositions of lysine. alpha. 380 and lysine. gamma. 486 in the acetylcholine receptor from Torpedo californica

Journal Article · Tue Apr 23 00:00:00 EDT 1991 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00230a041· OSTI ID:5510741

Dwyer, B P ^[1]

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1. Univ. of California, San Diego, La Jolla (USA)

The locations have been determined, with respect to the plasma membrane, of lysine {alpha}380 and lysine {gamma}486 in the {alpha} subunit and the {gamma} subunit, respectively, of the nicotinic acetylcholine receptor from Torpedo californica. Immunoadsorbents were constructed that recognize the carboxy terminus of the peptide GVKYIAE released by proteolytic digestion from positions 378-384 in the amino acid sequence of the {alpha} subunit of the acetylcholine receptor and the carboxy terminus of the peptide KYVP released by proteolytic digestion from positions 486-489 in the amino acid sequence of the {gamma} subunit. They were used to isolate these peptides from proteolytic digests of polypeptides from the acetylcholine receptor. Sealed vesicles containing the native acetylcholine receptor were labeled with pyridoxal phosphate and sodium ({sup 3}H)-borohydride. The effect of saponin on the incorporation of pyridoxamine phosphate into lysine {alpha}380 and lysine {gamma}486 from the acetylcholine receptor in these vesicles was assessed with the immunoadsorbents. The conclusions that follow from these results are that lysine {alpha}380 is on the inside surface of a vesicle and lysine {gamma}486 is on the outside surface. Because a majority (85%) of the total binding sites for {alpha}-bungarotoxin bind the toxin in the absence of saponin, the majority of the vesicles are right side out with the inside of the vesicle corresponding to the cytoplasmic surface and the outside of the vesicle corresponding to the extracytoplasmic, synaptic surface. Because lysine {alpha}380 and lysine {gamma}486 lie on opposite sides of the membrane, a membrane-spanning segment must be located between the two positions occupied by these two amino acids in the common sequence of a polypeptide of the acetylcholine receptor.

• OSTI ID: 5510741
• Journal Information: Biochemistry; (United States), Vol. 30:16; ISSN 0006-2960
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
LYSINE
TOPOLOGICAL MAPPING
RECEPTORS
MOLECULAR STRUCTURE
TOXINS
RADIORECEPTOR ASSAY
ACETYLCHOLINE
CELL MEMBRANES
GLYCOPROTEINS
NICOTINIC ACID
PYRIDOXINE
SAPONINS
TRITIUM COMPOUNDS
AMINES
AMINO ACIDS
AMMONIUM COMPOUNDS
ANTIGENS
AUTONOMIC NERVOUS SYSTEM AGENTS
AZINES
CARBOHYDRATES
CARBOXYLIC ACIDS
CELL CONSTITUENTS
DRUGS
ESTERS
GLYCOSIDES
HETEROCYCLIC ACIDS
HETEROCYCLIC COMPOUNDS
HYDROGEN COMPOUNDS
HYDROXY COMPOUNDS
ISOTOPE APPLICATIONS
MAPPING
MATERIALS
MEMBRANE PROTEINS
MEMBRANES
NEUROREGULATORS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PARASYMPATHOMIMETICS
PROTEINS
PYRIDINES
QUATERNARY COMPOUNDS
TOXIC MATERIALS
TRACER TECHNIQUES
TRANSFORMATIONS
VITAMIN B GROUP
VITAMINS
550201* - Biochemistry- Tracer Techniques