Recombinant pI(Cln) forms highly cation-selective channels when reconstituted into artificial and biological membranes
accesstype: All Open Access, Bronze, Green
has publication venue: Journal of General Physiology
most specific type: Article
page end: 736
most specific type: Document
origen: scopus
estado: published
e iscopus: 2-s2.0-0032217047
chemicalscas: Chloride Channels; Clns1a protein, rat; Ion Channels; Lipid Bilayers; Liposomes; Membranes, Artificial; Recombinant Proteins
doi: 10.1085/jgp.112.6.727
related by: authorship 3200
related by: authorship 3196
page start: 727
volume: 112
abstract: pI(Cln) has been proposed to be the swelling-activated anion channel responsible for I(Cl,swell), or a channel regulator. We tested the anion channel hypothesis by reconstituting recombinant pI(Cln) into artificial and biological membranes. Single channels were observed when pI(Cln) was reconstituted into planar lipid bilayers. In the presence of symmetrical 300 mM KCl, the channels had a high open probability and a slope conductance of 48 pS, and were outwardly rectifying. Reduction of trans KCl to 50 mM shifted the reversal potential by -31.2 ± 0.06 mV, demonstrating that the channel is at least seven times more selective for cations than for anions. Consistent with this finding, channel conductance was unaffected by substitution of Cl- with glutamate, but was undetectable when K%2b was replaced by N-methyl-d- glucamine. Reconstitution of pI(Cln) into liposomes increased 86Rb%2b uptake by three- to fourfold, but had no effect on 36Cl- uptake. Phosphorylation of pI(Cln) with casein kinase II or mutation of G54, G56, and G58 to alanine decreased channel open probability and 86Rb%2b uptake. When added to the external medium bathing Sf9 cells, pI(Cln) inserted into the plasma membrane and increased cell cation permeability. Taken together, these observations demonstrate that channel activity is due to pI(Cln) and not minor contaminant proteins. However, these findings do not support the hypothesis that pI(Cln) is the anion-selective I(Cl,swell) channel. The observed cation channel activity may reflect an as yet to be defined physiological function of pI(Cln), or may be a consequence of in vitro reconstitution of purified, recombinant protein.
type: Generically Dependent Continuant
related by: authorship 3201
type: Information Content Entity
freetext keyword: Cell volume; Liposomes; Planar lipid bilayer; Recombinant protein; Swelling-activated anion channels casein kinase II; cation channel; chloride ion; chlorine 36; glutamic acid; liposome; meglumine; membrane protein; potassium chloride; potassium ion; recombinant protein; rubidium 86; chloride channel; Clns1a protein, rat; ion channel; recombinant protein; animal cell; article; artificial membrane; cell membrane permeability; cell volume; controlled study; immunofluorescence microscopy; insect; ion conductance; light scattering; lipid bilayer; nonhuman; protein phosphorylation; animal; armyworm; cell line; cell membrane; cell membrane potential; chemistry; dog; genetics; in vitro study; metabolism; phosphorylation; rat; site directed mutagenesis; Animals; Cell Line; Cell Membrane; Cell Membrane Permeability; Chloride Channels; Dogs; Ion Channels; Lipid Bilayers; Liposomes; Membrane Potentials; Membranes, Artificial; Mutagenesis, Site-Directed; Phosphorylation; Rats; Recombinant Proteins; Spodoptera
has funding vehicle: National Institute of Diabetes and Digestive and Kidney Diseases, NIDDK: R01DK051610
type: Continuant
related by: authorship 3199
type: Document
type: Thing
has funding vehicle: National Institute of Neurological Disorders and Stroke, NINDS: R01NS030591
date time value: article date 668
related by: authorship 3197
crisid: 668
type: Entity
abstract: pI(Cln) has been proposed to be the swelling-activated anion channel responsible for I(Cl,swell), or a channel regulator. We tested the anion channel hypothesis by reconstituting recombinant pI(Cln) into artificial and biological membranes. Single channels were observed when pI(Cln) was reconstituted into planar lipid bilayers. In the presence of symmetrical 300 mM KCl, the channels had a high open probability and a slope conductance of 48 pS, and were outwardly rectifying. Reduction of trans KCl to 50 mM shifted the reversal potential by -31.2 ± 0.06 mV, demonstrating that the channel is at least seven times more selective for cations than for anions. Consistent with this finding, channel conductance was unaffected by substitution of Cl- with glutamate, but was undetectable when K was replaced by N-methyl-d- glucamine. Reconstitution of pI(Cln) into liposomes increased 86Rb uptake by three- to fourfold, but had no effect on 36Cl- uptake. Phosphorylation of pI(Cln) with casein kinase II or mutation of G54, G56, and G58 to alanine decreased channel open probability and 86Rb uptake. When added to the external medium bathing Sf9 cells, pI(Cln) inserted into the plasma membrane and increased cell cation permeability. Taken together, these observations demonstrate that channel activity is due to pI(Cln) and not minor contaminant proteins. However, these findings do not support the hypothesis that pI(Cln) is the anion-selective I(Cl,swell) channel. The observed cation channel activity may reflect an as yet to be defined physiological function of pI(Cln), or may be a consequence of in vitro reconstitution of purified, recombinant protein.
pmid: 9834142
related by: authorship 3202
related by: authorship 3198
type: Article
issue: 6
related by: authorship 3203

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Journal of General Physiology
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RDF description of Recombinant pI(Cln) forms highly cation-selective channels when reconstituted into artificial and biological membranes - http://orbis.uaslp.mx/vivo/individual/scopus667
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date: 2026-08-28T03:37:40
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National Institute of Diabetes and Digestive and Kidney Diseases, NIDDK: R01DK051610
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funding vehicle for: Recombinant pI(Cln) forms highly cation-selective channels when reconstituted into artificial and biological membranes

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National Institute of Neurological Disorders and Stroke, NINDS: R01NS030591
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