2FED

Structure of the E203Q mutant of the Cl-/H+ exchanger CLC-ec1 from E.Coli

Summary for 2FED

• Entry DOI: 10.2210/pdb2fed/pdb
• Related: 1OTS 2FEC 2FEE
• Descriptor: H(+)/Cl(-) exchange transporter clcA, Fab fragment, heavy chain, Fab fragment, light chain (3 entities in total)
• Functional Keywords: clc-ec1; clca_ecoli; chloride/proton exchange transporter, proton transport, membrane protein
• 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: 193138.37

Authors

Accardi, A.,Walden, M.P.,Nguitragool, W.,Jayaram, H.,Williams, C.,Miller, C. (deposition date: 2005-12-15, release date: 2006-01-03, Last modification date: 2024-10-30)

Primary citation

Accardi, A.,Walden, M.P.,Nguitragool, W.,Jayaram, H.,Williams, C.,Miller, C.
Separate ion pathways in a Cl-/H+ exchanger
J.Gen.Physiol., 126:563-570, 2005

PubMed Abstract: CLC-ec1 is a prokaryotic CLC-type Cl(-)/H+ exchange transporter. Little is known about the mechanism of H+ coupling to Cl-. A critical glutamate residue, E148, was previously shown to be required for Cl(-)/H+ exchange by mediating proton transfer between the protein and the extracellular solution. To test whether an analogous H+ acceptor exists near the intracellular side of the protein, we performed a mutagenesis scan of inward-facing carboxyl-bearing residues and identified E203 as the unique residue whose neutralization abolishes H+ coupling to Cl- transport. Glutamate at this position is strictly conserved in all known CLCs of the transporter subclass, while valine is always found here in CLC channels. The x-ray crystal structure of the E203Q mutant is similar to that of the wild-type protein. Cl- transport rate in E203Q is inhibited at neutral pH, and the double mutant, E148A/E203Q, shows maximal Cl- transport, independent of pH, as does the single mutant E148A. The results argue that substrate exchange by CLC-ec1 involves two separate but partially overlapping permeation pathways, one for Cl- and one for H+. These pathways are congruent from the protein's extracellular surface to E148, and they diverge beyond this point toward the intracellular side. This picture demands a transport mechanism fundamentally different from familiar alternating-access schemes.

PubMed: 16316975

Experimental method

X-RAY DIFFRACTION (3.317 Å)