High-level expression, functional reconstitution, and quaternary structure of a prokaryotic ClC-type chloride channel.
ClC-type anion-selective channels are widespread throughout eukaryotic organisms. BLAST homology searches reveal that many microbial genomes also contain members of the ClC family. An Escherichia coli-derived ClC Cl(-) channel homologue, "EriC," the product of the yadQ gene, was overexpressed in E. coli and purified in milligram quantities in a single-step procedure. Reconstitution of purified EriC into liposomes confers on these membranes permeability to anions with selectivity similar to that observed electrophysiologically in mammalian ClC channels. Cross-linking studies argue that EriC is a homodimer in both detergent micelles and reconstituted liposomes, a conclusion corroborated by gel filtration and analytical sedimentation experiments.
Maduke
M
M
Department of Biochemistry, Howard Hughes Medical Institute, Brandeis University, Waltham, Massachusetts 02254-9110, USA.
Pheasant
D J
DJ
Miller
C
C
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics; D018118:Chloride Channels / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D002713:Chlorine / Q000493:pharmacokinetics; D003432:Cross-Linking Reagents; D019281:Dimerization; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics*; D005404:Fixatives; D015964:Gene Expression Regulation, Bacterial; D005976:Glutaral; D008081:Liposomes / Q000737:chemistry; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000502:physiology; D020836:Protein Structure, Quaternary; D011868:Radioisotopes / Q000493:pharmacokinetics; D017386:Sequence Homology, Amino Acid; D014316:Tritium
D001426:Bacterial Proteins; D018118:Chloride Channels; D003432:Cross-Linking Reagents; D005404:Fixatives; D008081:Liposomes; D011868:Radioisotopes; D014316:Tritium; D002713:Chlorine; D005976:Glutaral