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Topology of an amiloride-binding protein.

Publication ,  Journal Article
Lin, C; Kieber-Emmons, T; Villalobos, AP; Foster, MH; Wahlgren, C; Kleyman, TR
Published in: J Biol Chem
January 28, 1994

Amiloride and structurally related compounds inhibit many transport proteins, enzymes, and drug or hormone receptors, although the topology of amiloride binding sites on these proteins has not been defined. We have previously raised and characterized a monoclonal antiamiloride antibody (mAb BA7.1) which is similar to epithelial Na+ channels in its specificity of binding of amiloride and amiloride analogs, suggesting that their amiloride binding sites may be similar in topology, mAb BA7.1 was used as a model system to analyze the three-dimensional conformation of an amiloride binding site. The photoactive amiloride analog 2'-methoxy-5'-nitrobenzamil specifically labeled the heavy chain of mAb BA7.1, suggesting that the heavy chain participates in amiloride binding. The nucleotide sequences of the variable regions of the heavy and light chains of mAb BA7.1 were determined and amino acid sequences deduced to analyze the structure of the amiloride binding site. A comparative modeling approach was used to construct a model of the amiloride binding domain of mAb BA7.1, and a docking procedure was used to place amiloride within this domain. The model indicated that planar aromatic amino acid resides form a pocket into which amiloride, a planar molecule, inserts. Constraints on amiloride binding predicted by this model correlated with the measured specificity of binding of amiloride analogs with mAb BA7.1. These results provide a potential guide for the identification of motifs or amino acid contact residues present within other amiloride-sensitive proteins.

Duke Scholars

Published In

J Biol Chem

ISSN

0021-9258

Publication Date

January 28, 1994

Volume

269

Issue

4

Start / End Page

2805 / 2813

Location

United States

Related Subject Headings

  • Sodium Channels
  • Sequence Homology, Amino Acid
  • Protein Conformation
  • Molecular Sequence Data
  • Models, Molecular
  • Mice
  • Immunoglobulin Light Chains
  • Immunoglobulin Heavy Chains
  • Enzyme-Linked Immunosorbent Assay
  • DNA Primers
 

Citation

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Lin, C., Kieber-Emmons, T., Villalobos, A. P., Foster, M. H., Wahlgren, C., & Kleyman, T. R. (1994). Topology of an amiloride-binding protein. J Biol Chem, 269(4), 2805–2813.
Lin, C., T. Kieber-Emmons, A. P. Villalobos, M. H. Foster, C. Wahlgren, and T. R. Kleyman. “Topology of an amiloride-binding protein.J Biol Chem 269, no. 4 (January 28, 1994): 2805–13.
Lin C, Kieber-Emmons T, Villalobos AP, Foster MH, Wahlgren C, Kleyman TR. Topology of an amiloride-binding protein. J Biol Chem. 1994 Jan 28;269(4):2805–13.
Lin, C., et al. “Topology of an amiloride-binding protein.J Biol Chem, vol. 269, no. 4, Jan. 1994, pp. 2805–13.
Lin C, Kieber-Emmons T, Villalobos AP, Foster MH, Wahlgren C, Kleyman TR. Topology of an amiloride-binding protein. J Biol Chem. 1994 Jan 28;269(4):2805–2813.

Published In

J Biol Chem

ISSN

0021-9258

Publication Date

January 28, 1994

Volume

269

Issue

4

Start / End Page

2805 / 2813

Location

United States

Related Subject Headings

  • Sodium Channels
  • Sequence Homology, Amino Acid
  • Protein Conformation
  • Molecular Sequence Data
  • Models, Molecular
  • Mice
  • Immunoglobulin Light Chains
  • Immunoglobulin Heavy Chains
  • Enzyme-Linked Immunosorbent Assay
  • DNA Primers