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Crystal structure of the E148D mutant CLC-ec1 in 50 mM bromide
Structural highlights
FunctionCLCA_ECOLI Proton-coupled chloride transporter. Functions as antiport system and exchanges two chloride ions for 1 proton. Probably acts as an electrical shunt for an outwardly-directed proton pump that is linked to amino acid decarboxylation, as part of the extreme acid resistance (XAR) response.[1] [2] [3] [4] [5] Publication Abstract from PubMedThe CLC family of proteins are involved in a variety of physiological processes to control cellular chloride concentration. Two distinct classes of CLC proteins, Cl(-) channels and Cl(-)/H(+) antiporters, have been functionally and structurally investigated over the last several decades. Previous studies have suggested that the conformational heterogeneity of the critical glutamate residue, Gluex, could explain the transport cycle of CLC-type Cl(-)/H(+) antiporters. However, the presence of multiple conformations (Up, Middle, and Down) of the Gluex has been suggested from combined structural snapshots of 2 different CLC antiporters: CLC-ec1 from Escherichia coli and cmCLC from a thermophilic red alga, Cyanidioschyzon merolae Thus, we aimed to investigate further the heterogeneity of Gluex-conformations in CLC-ec1, the most deeply studied CLC antiporter, at both functional and structural levels. Here, we show that the crystal structures of the Gluex mutant E148D and wild-type CLC-ec1 with varying anion concentrations suggest a structural intermediate, the "Midlow" conformation. We also found that an extra anion can be located above the external Cl(-)-binding site in the E148D mutant when the anion concentration is high. Moreover, we observed that a carboxylate in solution can occupy either the external or central Cl(-)-binding site in the ungated E148A mutant using an anomalously detectable short carboxylic acid, bromoacetate. These results lend credibility to the idea that the Gluex can take at least 3 distinct conformational states during the transport cycle of a single CLC antiporter. Mutation of external glutamate residue reveals a new intermediate transport state and anion binding site in a CLC Cl(-)/H(+) antiporter.,Park K, Lee BC, Lim HH Proc Natl Acad Sci U S A. 2019 Aug 13. pii: 1901822116. doi:, 10.1073/pnas.1901822116. PMID:31409705[6] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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