2HTK

Structure of the Escherichia coli ClC chloride channel Y445A mutant and Fab complex

Summary for 2HTK

• Entry DOI: 10.2210/pdb2htk/pdb
• Related: 1OTS 2HLF 2HT2 2HT3 2HT4 2HTL
• Descriptor: H(+)/Cl(-) exchange transporter clcA, Fab fragment, Heavy chain, Fab fragment, Light chain, ... (4 entities in total)
• Functional Keywords: clc family of channel and transporters, h+/cl- antiporter, membrane protein, fab complex
• Biological source: Escherichia coli; More
• Cellular location: Cell inner membrane; Multi-pass membrane protein (Probable): P37019
• Total number of polymer chains: 6
• Total formula weight: 194321.15

Authors

Accardi, A.,Lobet, S.,Williams, C.,Miller, C.,Dutzler, R. (deposition date: 2006-07-26, release date: 2006-09-19, Last modification date: 2024-10-16)

Primary citation

Accardi, A.,Lobet, S.,Williams, C.,Miller, C.,Dutzler, R.
Synergism between halide binding and proton transport in a CLC-type exchanger
J.Mol.Biol., 362:691-699, 2006

PubMed 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.

PubMed: 16949616
DOI: 10.1016/j.jmb.2006.07.081

Experimental method

X-RAY DIFFRACTION (3.41 Å)