Many proteins of the CLC gene family are Cl(-) channels, whereas others, like the bacterial ecClC-1 or mammalian ClC-4 and -5, mediate Cl(-)/H(+) exchange. Mutating a "gating glutamate" (Glu-224 in ClC-4 and Glu-211 in ClC-5) converted these exchangers into anion conductances, as did the neutralization of another, intracellular "proton glutamate" in ecClC-1. We show here that neutralizing the proton glutamate of ClC-4 (Glu-281) and ClC-5 (Glu-268), but not replacing it with aspartate, histidine, or tyrosine, rather abolished Cl(-) and H(+) transport. Surface expression was unchanged by these mutations. Uncoupled Cl(-) transport could be restored in the ClC-4(E281A) and ClC-5(E268A) proton glutamate mutations by additionally neutralizing the gating glutamates, suggesting that wild type proteins transport anions only when protons are supplied through a cytoplasmic H(+) donor. Each monomeric unit of the dimeric protein was found to be able to carry out Cl(-)/H(+) exchange independently from the transport activity of the neighboring subunit. NO(3)(-) or SCN(-) transport was partially uncoupled from H(+) countertransport but still depended on the proton glutamate. Inserting proton glutamates into CLC channels altered their gating but failed to convert them into Cl(-)/H(+) exchangers. Noise analysis indicated that ClC-5 switches between silent and transporting states with an apparent unitary conductance of 0.5 picosiemens. Our results are consistent with the idea that Cl(-)/H(+) exchange of the endosomal ClC-4 and -5 proteins relies on proton delivery from an intracellular titratable residue at position 268 (numbering of ClC-5) and that the strong rectification of currents arises from the voltage-dependent proton transfer from Glu-268 to Glu-211.
Enables the active transport of a solute across a membrane by a mechanism whereby two or more species are transported in opposite directions in a tightly coupled process not directly linked to a form of energy other than chemiosmotic energy. The reaction is: solute A(out) + solute B(in) = solute A(in) + solute B(out).
Catalysis of facilitated diffusion of a chloride (by an energy-independent process) involving passage through a transmembrane aqueous pore or channel without evidence for a carrier-mediated mechanism.
Am. J. Physiol. 277, C948-54-C948-54 (1999)[PubMed:10564087]
ClC-4 gene was isolated as a putative Cl(-) channel. Due to a lack of functional expression of ClC-4, its physiological role remains unknown. We isolated a human ClC-4 clone (hClC-4sk) from human skeletal muscles and stably transfected it to Chinese hamster ovary cells. Whole cell patch-clamp studies showed that the hClC-4sk channel was activated by external acidic pH and inhibited by DIDS. It passed a strong outward Cl(-) current with a permeability sequence of I(-) > Cl(-) > F(-). The hClC-4sk has consensus sites for phosphorylation by protein kinase A (PKA); however, stimulation of PKA had no effect on the currents. hClC-4sk mRNA was expressed in excitable tissues, such as heart, brain, and skeletal muscle. These functional characteristics of hClC-4sk provide a clue to its physiological role in excitable cells.
Catalysis of the transmembrane transfer of a chloride ion by a voltage-gated channel. A voltage-gated channel is a channel whose open state is dependent on the voltage across the membrane in which it is embedded.
Am. J. Physiol. 277, C948-54-C948-54 (1999)[PubMed:10564087]
ClC-4 gene was isolated as a putative Cl(-) channel. Due to a lack of functional expression of ClC-4, its physiological role remains unknown. We isolated a human ClC-4 clone (hClC-4sk) from human skeletal muscles and stably transfected it to Chinese hamster ovary cells. Whole cell patch-clamp studies showed that the hClC-4sk channel was activated by external acidic pH and inhibited by DIDS. It passed a strong outward Cl(-) current with a permeability sequence of I(-) > Cl(-) > F(-). The hClC-4sk has consensus sites for phosphorylation by protein kinase A (PKA); however, stimulation of PKA had no effect on the currents. hClC-4sk mRNA was expressed in excitable tissues, such as heart, brain, and skeletal muscle. These functional characteristics of hClC-4sk provide a clue to its physiological role in excitable cells.
The directed movement of substances (such as macromolecules, small molecules, ions) into, out of or within a cell, or between cells, or within a multicellular organism by means of some agent such as a transporter or pore.
In the framework of constructing a comprehensive transcript map of the human Xp22.3 region, we identified an evolutionary conserved CpG island and cloned the corresponding gene. The predicted 760 amino acid protein encoded by this gene contains 12 hydrophobic domains and shares significant sequence and structural similarities with all the previously isolated members of a recently identified family of voltage-gated chloride channels (the 'CIC family'). This gene, termed CICN4 (Chloride Channel 4), contains at least 10 exons spanning 60 to 80 kb on the X chromosome. In contrast to most genes isolated from the human Xp22.3 region, the CICN4 gene does not share homology with the Y chromosome and it is conserved in mouse and hamster. Expression studies revealed the presence of a 7.5 kb transcript which is particularly abundant in skeletal muscle and is also detectable in brain and heart. These data suggest that we have identified a new voltage-gated chloride channel which is encoded by a gene located in the distal short arm of the X chromosome.
The CLC channel family contains both chloride channels and proton-coupled anion transporters that exchange chloride or another anion for protons. The presence of conserved gating glutamate residues is typical for family members that function as antiporters.
Protein involved in the transport of a solute across a biological membrane coupled, directly, to the transport of a different solute in the opposite direction.
Protein involved in the transport of ions. Such proteins are usually transmembrane and mediate a movement of ions across cell membranes. Transport may be passive (facilitated diffusion; down the electrochemical gradient), or active (against the electrochemical gradient). Active transport requires energy which may come from light, oxidation reactions, ATP hydrolysis, or cotransport of other ions or molecules.
Protein involved in the transport of a molecule (metabolite, protein, etc), a ion or an electron across cell membranes, inside the cell or in a tissue fluid.
A reference proteome is a set of protein sequences derived from a complete proteome which constitutes a defined standard for a particular user community. Reference proteomes are manually defined according to a number of criteria. They cover the proteomes of well- studied model organisms and other proteomes of interest for biomedical and biotechnological research. Reference proteomes have been selected to provide broad coverage of the tree of life, and constitute a representative cross-section of the taxonomic diversity to be found within UniProtKB.
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