Site of Covalent Labeling by a Photoreactive Batrachotoxin Derivative near Transmembrane Segment IS6 of the Sodium Channel Subunit (*)

The binding site for batrachotoxin, a lipid-soluble neurotoxin acting at Na channel receptor site 2, was localized using a photoreactive, radiolabeled batrachotoxin derivative to covalently label purified and reconstituted rat brain Na channels. In the presence of the brevetoxin 1 from Ptychodiscus brevis and the pyrethroid RU51049, positive allosteric enhancers of batrachotoxin binding, a protein with an apparent molecular mass of 240 kDa corresponding to the Na channel α subunit was specifically covalently labeled. The region of the α subunit specifically photolabeled by the photoreactive batrachotoxin derivative was identified by antibody mapping of proteolytic fragments. Even after extensive trypsinization, an anti-peptide antibody recognizing an amino acid sequence adjacent to Na channel transmembrane segment IS6 was able to immunoprecipitate up to 70% of the labeled peptides. Analysis of a more complete digestion with trypsin or V8 protease indicated that the batrachotoxin receptor site is formed in part by a portion of domain I. The identification of a specifically immunoprecipitated photolabeled 7.3-kDa peptide containing transmembrane segment S6 from domain I restricted the site of labeling to residues Asn-388 to Glu-429 if V8 protease digestion was complete or Leu-380 to Glu-429 if digestion was incomplete. These results implicate the S6 transmembrane region of domain I of the Na channel α subunit as an important component of the batrachotoxin receptor site.

• Publication year

  1996

• Venue

  Journal of Biological Chemistry

• Publication date

  1996-05-10

• Fields of study

  Biology, Medicine, Chemistry

• Identifiers
  DOI 10.1074/JBC.271.19.11261PMID 8626676
• External record

  Open on Semantic Scholar

• Source metadata

  Semantic Scholar, PubMed

• No claims are published for this paper.

• No concepts are published for this paper.

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