We are seeing patients in-person and through Video Visits. Learn more about how we’re keeping you safe and please review our updated visitor policy. Please also consider supporting Weill Cornell Medicine’s efforts to support our front-line workers.
Department of Anesthesiology

You are here

Synergism between halide binding and proton transport in a CLC-type exchanger.

TitleSynergism between halide binding and proton transport in a CLC-type exchanger.
Publication TypeJournal Article
Year of Publication2006
AuthorsAccardi A, Lobet S, Williams C, Miller C, Dutzler R
JournalJ Mol Biol
Volume362
Issue4
Pagination691-9
Date Published2006 Sep 29
ISSN0022-2836
KeywordsBromides, Chloride Channels, Escherichia coli, Escherichia coli Proteins, Ion Transport, Mutant Proteins, Protein Structure, Secondary, Protons
Abstract

The Cl-/H+ exchange-transporter CLC-ec1 mediates stoichiometric transmembrane exchange of two Cl- ions for one proton. A conserved tyrosine residue, Y445, coordinates one of the bound Cl- ions visible in the structure of this protein and is located near the intersection of the Cl- and H+ pathways. Mutants of this tyrosine were scrutinized for effects on the coupled transport of Cl- and H+ determined electrophysiologically and on protein structure determined crystallographically. Despite the strong conservation of Y445 in the CLC family, substitution of F or W at this position preserves wild-type transport behavior. Substitution by A, E, or H, however, produces uncoupled proteins with robust Cl- transport but greatly impaired movement of H+. The obligatory 2 Cl-/1 H+ stoichiometry is thus lost in these mutants. The structures of all the mutants are essentially identical to wild-type, but apparent anion occupancy in the Cl- binding region correlates with functional H+ coupling. In particular, as determined by anomalous diffraction in crystals grown in Br-, an electrophysiologically competent Cl- analogue, the well-coupled transporters show strong Br- electron density at the "inner" and "central" Cl- binding sites. However, in the uncoupled mutants, Br- density is absent at the central site, while still present at the inner site. An additional mutant, Y445L, is intermediate in both functional and structural features. This mutant clearly exchanges H+ for Cl-, but at a reduced H+-to-Cl- ratio; likewise, both the central and inner sites are occupied by Br-, but the central site shows lower Br- density than in wild-type (or in Y445F,W). The correlation between proton coupling and central-site occupancy argues that halide binding to the central transport site somehow facilitates movement of H+, a synergism that is not readily understood in terms of alternating-site antiport schemes.

DOI10.1016/j.jmb.2006.07.081
Alternate JournalJ. Mol. Biol.
PubMed ID16949616
Grant ListGM-31768 / GM / NIGMS NIH HHS / United States