Antiidiotypic antibody recognizes an amiloride binding domain within the alpha subunit of the epithelial Na+ channel.

Published

Journal Article

We previously raised an antibody (RA6.3) by an antiidiotypic approach which was designed to be directed against an amiloride binding domain on the epithelial Na+ channel (ENaC). This antibody mimicked amiloride in that it inhibited transepithelial Na+ transport across A6 cell monolayers. RA6.3 recognized a 72-kDa polypeptide in A6 epithelia treated with tunicamycin, consistent with the size of nonglycosylated Xenopus laevis alphaENaC. RA6.3 specifically recognized an amiloride binding domain within the alpha-subunit of mouse and bovine ENaC. The deduced amino acid sequence of RA6.3 was used to generate a three-dimensional model structure of the antibody. The combining site of RA6.3 was epitope mapped using a novel computer-based strategy. Organic residues that potentially interact with the RA6.3 combining site were identified by data base screening using the program LUDI. Selected residues docked to the antibody in a manner corresponding to the ordered linear array of amino acid residues within an amiloride binding domain on the alpha-subunit of ENaC. A synthetic peptide spanning this domain inhibited the binding of RA6.3 to alphaENaC. This analysis provided a novel approach to develop models of antibody-antigen interaction as well as a molecular perspective of RA6.3 binding to an amiloride binding domain within alphaENaC.

Full Text

Duke Authors

Cited Authors

  • Kieber-Emmons, T; Lin, C; Foster, MH; Kleyman, TR

Published Date

  • April 2, 1999

Published In

Volume / Issue

  • 274 / 14

Start / End Page

  • 9648 - 9655

PubMed ID

  • 10092651

Pubmed Central ID

  • 10092651

International Standard Serial Number (ISSN)

  • 0021-9258

Digital Object Identifier (DOI)

  • 10.1074/jbc.274.14.9648

Language

  • eng

Conference Location

  • United States