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Atp induces conformational changes in the carboxyl-terminal region of clc-5

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Overview

authors

  • Wellhauser, L.
  • Luna-Chavez, C.
  • D'Antonio, C.
  • Tainer, John
  • Bear, C. E.

publication date

  • February 2011

journal

  • Journal of Biological Chemistry  Journal

abstract

  • ATP binding enhances the activity of ClC-5, the transporter mutated in Dent disease, a disease affecting the renal proximal tubule. Previously, the ATP binding site was revealed in x-ray crystal structures of the cytoplasmic region of this membrane protein. Disruption of this site by mutagenesis (Y617A-ClC-5) reduced the functional expression and ATP-dependent regulation of the full-length transporter in Xenopus oocytes. However, insight into the conformational changes underlying ATP-dependent regulation is lacking. Here, we show that ATP binding induces a change in protein conformation. Specifically, small angle x-ray scattering experiments indicate that ATP binding promotes a clamp-like closure of the isolated ClC-5 carboxyl-terminal region. Limited proteolysis studies show that ATP binding induces conformational compaction of the carboxyl-terminal region in the intact membrane protein as well. In the context of fibroblasts and proximal tubule epithelial cells, disruption of the ATP binding site in full-length ClC-5 (Y617A-ClC-5) led to a defect in processing and trafficking out of the endoplasmic reticulum. These latter findings account for the decrease in functional expression previously reported for this ATP-binding mutant and prompt future study of a model whereby conformational compaction caused by ATP binding promotes biosynthetic maturation.

subject areas

  • Adenosine Triphosphate
  • Amino Acid Substitution
  • Animals
  • Binding Sites
  • CHO Cells
  • Cricetinae
  • Cricetulus
  • Dent Disease
  • Endoplasmic Reticulum
  • Humans
  • Mutagenesis
  • Mutation, Missense
  • Oocytes
  • Peptide Mapping
  • Protein Structure, Tertiary
  • Protein Transport
  • Repressor Proteins
  • Xenopus laevis
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Identity

PubMed Central ID

  • PMC3057859

International Standard Serial Number (ISSN)

  • 0021-9258

Digital Object Identifier (DOI)

  • 10.1074/jbc.M110.175877

PubMed ID

  • 21173145
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Additional Document Info

start page

  • 6733

end page

  • 6741

volume

  • 286

issue

  • 8

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