Wyniki wyszukiwania

1

Transmembrane segment M2 of glycine receptor as a model system for the pore-forming structure of ion channels

100%

Bednarczyk P., Szewczyk A., Dolowy K.

Acta Biochimica Polonica

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2002

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tom 49

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nr 4

The glycine receptor belongs to the ligand-gated ion channel superfamily. It is a chloride conducting channel composed of four transmembrane domains. It was previously shown that the second transmembrane domain (M2) of the glycine receptor forms an ion conduction pathway throught lipid bilayers. The amino-acid sequence of the transmembrane segment M2 of the glycine receptor has a high homology to all receptors of the ligand-gated ion channel superfamily. In our report, we have used a synthetic M2 peptide. It was incorporated into a planar membrane of known lipid composition and currents induced by M2 were measured by the Black Lipid Membrane technique. When the planar lipid bilayer was composed of 75% phosphatidylethanolamine and 25% phosphatidylserine, the reversal potential measured in a 150/600 mM KCl (cis/trans) gradient was -19 mV suggesting that the examined pore was preferential to anions, Pk/Pci = 0.25. In contrast, when 75% phosphatidylserine and 25% phosphatidylethanolamine was used, the reversal potential was +20 mV and the pore was preferential to cations, Pk/Pci = 4.36. Single-channel currents were recorded with two predominant amplitudes corresponding to the main-conductance and sub-conductance states. Both conductance states (about 12 pS and 30 pS) were measured in a symmetric solution of 50 mM KCl. The observed single-channel properties suggest that the selectivity and conductance of the pore formed by the M2 peptide of the glycine receptor depend on the lipid composition of the planar bilayer.

2

The heterogeneity of ion channels in chromaffin granule membranes

100%

Hordejuk R., Szewczyk A., Dolowy K.

Cellular and Molecular Biology Letters

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2006

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tom 11

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nr 3

Chromaffin granules are involved in catecholamine synthesis and traffic in the adrenal glands. The transporting membrane proteins of chromaffin granules play an important role in the ion homeostasis of these organelles. In this study, we characterized components of the electrogenic 86Rb+ flux observed in isolated chromaffin granules. In order to study single channel activity, chromaffin granules from the bovine adrenal medulla were incorporated into planar lipid bilayers. Four types of cationic channel were found, each with a different conductance. The unitary conductances of the potassium channels are 360 ± 10 pS, 220 ± 8 pS, 152 ± 8 pS and 13 ± 3 pS in a gradient of 450/150 mM KCl, pH 7.0. A multiconductance potassium channel with a conductivity of 110 ± 8 pS and 31 ± 4 pS was also found. With the exception of the 13 pS conductance channel, all are activated by depolarizing voltages. One type of chloride channel was also found. It has a unitary conductance of about 250 pS in a gradient of 500/150 mM KCl, pH 7.0.

3

The prion peptide forms ion channels in planar lipid bilayers

100%

Berest V., Rutkowski M., Rolka K., Legowska A., Debska G., Stepkowski D., Szewczyk A.

Cellular and Molecular Biology Letters

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2003

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tom 08

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nr 2

One of the hypotheses concerning the pathogenic properties of the prion protein, considers its influence on cellular ion homeostasis. Using the lipid bilayer technique, the influence of prion-derived peptides on the lipid bilayer conductance was characterized. To evaluate the physiological significance and possible pathological functions of the peptides, their effect on the membrane potential and respiration rate of hippocampal mitochondria was also studied. We used a peptide bearing the human prion protein sequence YSNQNNF (PrP [169- 175]), and peptide SSQNNF (PrP [170-175]) bearing a naturally-occurring mutation in position 171 [N→S] linked to schizoaffective diseases in humans (Samaia, H.B., Mari, J.J., Vallada, H.P., Moura R.P., Simpson A.J.G., Brentani R.R. A prion-linked psychiatric disorder. Nature 390 (1997) 241). In this report, we show that PrP [170-175] N171S increases the conductance of planar lipid bilayers. Based on the conductance of single channel currents recorded in 500/500 mM KCl (cis/trans), we found a single channel conductance of 8 to 26 pS. The native prion peptide PrP [169-175] does not form ion channels in the lipid bilayer. Neither of the peptides significantly changed the membrane potential or respiration rate of isolated rat hippocampal mitochondria. We propose a possible mechanism for channel formation by aggregation of the prion-derived peptide.

4

Stilbene derivatives inhibit the activity of the inner mitochondrial membrane chloride channels

80%

Koszela-Piotrowska I., Choma K., Bednarczyk P., Dolowy K., Szewczyk A., Kunz W. S., Malekova L., Kominkova V., Ondrias K.

Cellular and Molecular Biology Letters

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2007

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tom 12

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nr 4

493-508

Ion channels selective for chloride ions are present in all biological membranes, where they regulate the cell volume or membrane potential. Various chloride channels from mitochondrial membranes have been described in recent years. The aim of our study was to characterize the effect of stilbene derivatives on single-chloride channel activity in the inner mitochondrial membrane. The measurements were performed after the reconstitution into a planar lipid bilayer of the inner mitochondrial membranes from rat skeletal muscle (SMM), rat brain (BM) and heart (HM) mitochondria. After incorporation in a symmetric 450/450 mM KCl solution (cis/trans), the chloride channels were recorded with a mean conductance of 155 ± 5 pS (rat skeletal muscle) and 120 ± 16 pS (rat brain). The conductances of the chloride channels from the rat heart mitochondria in 250/50 mM KCl (cis/trans) gradient solutions were within the 70–130 pS range. The chloride channels were inhibited by these two stilbene derivatives: 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) and 4-acetamido-4′-isothiocyanostilbene-2,2′-disulfonic acid (SITS). The skeletal muscle mitochondrial chloride channel was blocked after the addition of 1 mM DIDS or SITS, whereas the brain mitochondrial channel was blocked by 300 μM DIDS or SITS. The chloride channel from the rat heart mitochondria was inhibited by 50–100 μM DIDS. The inhibitory effect of DIDS was irreversible. Our results confirm the presence of chloride channels sensitive to stilbene derivatives in the inner mitochondrial membrane from rat skeletal muscle, brain and heart cells.

Ograniczanie wyników

3 Cellular and Molecular Biology Letters

1 Acta Biochimica Polonica

4 Szewczyk A.

3 Dolowy K.

2 Bednarczyk P.

1 Berest V.

1 Choma K.

1 Debska G.

1 Hordejuk R.

1 Kominkova V.

1 Koszela-Piotrowska I.

1 Kunz W. S.

1 Legowska A.

1 Malekova L.

1 Ondrias K.

1 Rolka K.

1 Rutkowski M.

1 Stepkowski D.

1 2007

1 2006

1 2003

1 2002

Transmembrane segment M2 of glycine receptor as a model system for the pore-forming structure of ion channels

The heterogeneity of ion channels in chromaffin granule membranes

The prion peptide forms ion channels in planar lipid bilayers

Stilbene derivatives inhibit the activity of the inner mitochondrial membrane chloride channels