• Genomics & Informatics
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• v.21 no.3
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• pp.41.1-41.11
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• 2023

The Dengue virus M protein is a 75 amino acid polypeptide with two helical transmembranes (TM). The TM domain oligomerizes to form an ion channel, facilitating viral release from the host cells. The M protein has a critical role in the virus entry and life cycle, making it a potent drug target. The oligomerization of the monomeric protein was studied using ab initio modeling and molecular dynamics simulation in an implicit membrane environment. The representative structures obtained showed pentamer as the most stable oligomeric state, resembling an ion channel. Glutamic acid, threonine, serine, tryptophan, alanine, isoleucine form the pore-lining residues of the pentameric channel, conferring an overall negative charge to the channel with approximate length of 51.9 Å. Residue interaction analysis for M protein shows that Ala94, Leu95, Ser112, Glu124, and Phe155 are the central hub residues representing the physicochemical interactions between domains. The virtual screening with 165 different ion channel inhibitors from the ion channel library shows monovalent ion channel blockers, namely lumacaftor, glipizide, gliquidone, glisoxepide, and azelnidipine to be the inhibitors with high docking scores. Understanding the three-dimensional structure of M protein will help design therapeutics and vaccines for Dengue infection.

• Biomedical Science Letters
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• v.18 no.2
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• pp.96-103
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• 2012

Kv 4.2 ion channel protein has an ability to open at subthreshold membrane potentials and to recover quickly from inactivation. That is very important for neuronal signal transmission in vertebrate brain. In order to purify Kv 4.2 protein, the novel purification methods were experimented. The purification procedure utilized chromatography on DE-52 ion exchange column and affinity chromatography on a WGA-Sepharose 4B, and Kv 4.2 affinity column chromatography. It was found that 0.5% (wt./vol.) Triton X-100 detergent in lysis buffer worked well for Kv 4.2 protein solubilization from rat brain membrane. Protein quantitative determination was conducted by BCA method at 562 nm for each purification step to avoid determination interference of protein at 280 nm by detergent. The confirmation of Kv 4.2 existence and amount is performed using by SDS-PAGE/immunoblotting or 96-well dot blotting. The Kv 4.2 without interacting protein that contains carbohydrate, was purified from novel biochemical 3-steps purification method for further research.

• BMB Reports
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• v.55 no.11
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• pp.528-534
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• 2022

Mitochondria are cellular organelles that perform various functions within cells. They are responsible for ATP production, cell-signal regulation, autophagy, and cell apoptosis. Because the mitochondrial proteins that perform these functions need Ca²⁺ ions for their activity, mitochondria have ion channels to selectively uptake Ca²⁺ ions from the cytoplasm. The ion channel known to play the most important role in the Ca²⁺ uptake in mitochondria is the mitochondrial calcium uniporter (MCU) holo-complex located in the inner mitochondrial membrane (IMM). This ion channel complex exists in the form of a complex consisting of the pore-forming protein through which the Ca²⁺ ions are transported into the mitochondrial matrix, and the auxiliary protein involved in regulating the activity of the Ca²⁺ uptake by the MCU holo-complex. Studies of this MCU holo-complex have long been conducted, but we didn't know in detail how mitochondria uptake Ca²⁺ ions through this ion channel complex or how the activity of this ion channel complex is regulated. Recently, the protein structure of the MCU holo-complex was identified, enabling the mechanism of Ca²⁺ uptake and its regulation by the MCU holo-complex to be confirmed. In this review, I will introduce the mechanism of action of the MCU holo-complex at the molecular level based on the Cryo-EM structure of the MCU holo-complex to help understand how mitochondria uptake the necessary Ca²⁺ ions through the MCU holo-complex and how these Ca²⁺ uptake mechanisms are regulated.

• ALGAE
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• v.36 no.4
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• pp.315-326
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• 2021

Ion channels are membrane protein complexes mediating passive ion flux across the cell membranes. Every organism has a certain set of ion channels that define its physiology. Dinoflagellates are ecologically important microorganisms characterized by effective physiological adaptability, which backs up their massive proliferations that often result in harmful blooms (red tides). In this study, we used a bioinformatics approach to identify homologs of known ion channels that belong to 36 ion channel families. We demonstrated that the versatility of the dinoflagellate physiology is underpinned by a high diversity of ion channels including homologs of animal and plant proteins, as well as channels unique to protists. The analysis of 27 transcriptomes allowed reconstructing a consensus ion channel repertoire (channelome) of dinoflagellates including the members of 31 ion channel families: inwardly-rectifying potassium channels, two-pore domain potassium channels, voltage-gated potassium channels (K_v), tandem K_v, cyclic nucleotide-binding domain-containing channels (CNBD), tandem CNBD, eukaryotic ionotropic glutamate receptors, large-conductance calcium-activated potassium channels, intermediate/small-conductance calcium-activated potassium channels, eukaryotic single-domain voltage-gated cation channels, transient receptor potential channels, two-pore domain calcium channels, four-domain voltage-gated cation channels, cation and anion Cys-loop receptors, small-conductivity mechanosensitive channels, large-conductivity mechanosensitive channels, voltage-gated proton channels, inositole-1,4,5-trisphosphate receptors, slow anion channels, aluminum-activated malate transporters and quick anion channels, mitochondrial calcium uniporters, voltage-dependent anion channels, vesicular chloride channels, ionotropic purinergic receptors, animal volage-insensitive cation channels, channelrhodopsins, bestrophins, voltage-gated chloride channels H⁺/Cl^- exchangers, plant calcium-permeable mechanosensitive channels, and trimeric intracellular cation channels. Overall, dinoflagellates represent cells able to respond to physical and chemical stimuli utilizing a wide range of G-protein coupled receptors- and Ca²⁺-dependent signaling pathways. The applied approach not only shed light on the ion channel set in dinoflagellates, but also provided the information on possible molecular mechanisms underlying vital cellular processes dependent on the ion transport.

• Journal of Embryo Transfer
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• v.30 no.4
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• pp.277-282
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• 2015

Cryopreservation affects osmotic tolerance and intracellular ion concentration through changes in expression levels of water and ion channels. Control of these changes is important for cell survival after cryopreservation. Relatively little is known about changes in $K^+$ channel expression compared to water channel expression. This study was performed to investigate changes in TASK-2 channel (KCNK5: potassium channel, subfamily K, member 5), a member of two-pore domain $K^+$ channel family, in cryopreserved mouse ovaries. Cryopreservation increased TASK-2 mRNA expression in mouse ovaries. In addition, TASK-2 protein expression was upregulated in vitrified and slowly frozen ovaries. TASK-2 protein was expressed in all area of granulosa cells that surround the oocyte within the follicle, except nucleus. Viability of cells overexpressed with TASK-2 was higher than that of vector-transfected cells. Our results found that TASK-2 expression was increased by cryopreservation and overexpression of TASK-2 decreased cryopreservation-induced cell death. These results suggest that TASK-2 upregulation might reduce cryodamage.

• The Korean Journal of Pharmacology
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• v.31 no.3
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• pp.355-363
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• 1995

It has been well known that the intracellular calcium concentration $([Ca^{2+}]_i)$ in living cell is very sensitive to live or to survive, but the transmembrane system of calcium ion, especially mechanism of calcium ion movement in unexcitable state has been little elucidated. Though many proposed theories for calcium ion transport have been reported, it is still unclear that how could the sustained maintenance in cytosolic calcium level be done in cell. Since one of possible mechanisms of calcium transport may be related to the acetylcholine receptor-linked calcium channel, author performed experiment to elucidate this mechanism of calcium influx related to cholinergic receptor in ml muscarinic receptor-transfected RBL-2H3 cell-line. 1) The effects of carbachol both on calcium ion influx and on the secretion of hexosaminidase were respectively observed in the manner of time-related or concentration-dependent pattern in this model. 2) The effects of several metal cations on calcium transport were shown in carbachol-induced cell-line. 3) Atropine was administered to examine the relationship between cholinergic receptor and calcium ion influx in this model. 4) PMA (Phorbol 12-myristate 13-acetate) or PTx (Pertussis toxin) was respectively administered to examine the secondary mediator which involved pathway of calcium ion movement in carbachol-induced cell-line. The results of this experiments were as follows; 1) Carbachol significantly stimulated both the calcium influx and the secretion of hexosaminidase in the manner of the concentration-dependent pattern. 2) Atropine potently blocked the effects of carbachol in concentration-response manner. 3) Administered metal cations inhibited the calcium influx in carbachol-stimulated this model to the concentration-related pattern. 4) PMA did not inhibit carbachol-induced secretion of hexosaminidase, but blocked the calcium influx in this cell-line. 5) The suppression of carbachol-induced hexosaminidase secretion was shown in PTx-treated cell -line.

• Journal of Ginseng Research
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• v.20 no.3
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• pp.274-283
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• 1996

In this study, the effects of red ginseng components [ginsenosides (total saponins and $Rg_1$) on the function of ryanodine receptor (RyR) -$Ca^{2+}$ release channel complex protein (named as RyR or $Ca^{2+}$ channel), a membrane protein in sarcoplasmic reticulum (SR) of rabbit skeletal muscle were examined at the SR vesicle's level and the molecular levels with Chaps-solubilized and purified $Ca^{2+}$ channel protein and with reconstituted proteoliposomes by dialysis. The results were as follows. 1. The binding of ryanodine known as inhibitor of muscle contraction to the RyR was decreased at the whole range of concentration ($10^2$~$10^7$%) by these two ginseng components. In heavy SR vesicles, Chaps-solubilized and purified $Ca^{2+}$ channel protein, and reconstituted vesicles, its maximal inhibition by total saponins was shown at the concentration of $10^3$, $10^3$%, and $10^5$% respectively, and by gin- senoside $Rg_1}$) each was $10^3$%, $10^3$%, and $10^4$%. 2. The release of $Ca^{2+}$ ion through $Ca^{2+}$ channel in heavy SR vesicles and reconstituted proteoliposomes was increased as a whole by these two ginseng components, and particularly maximal release by both of them was shown at the range of $10^4$~$10^6$%. These results were seemed to be caused by conformational change of $Ca^{2+}$ release channel protein (RyR) by red ginseng components [ginsenosides (total saponins and $Rg_1}$).

• BMB Reports
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• v.44 no.9
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• pp.559-565
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• 2011

Protein kinase C (PKC) is a central enzyme that modulates numerous biological functions. For this reason, specific PKC inhibitors/activators are required to study PKC-related signaling mechanisms. To date, although many PKC inhibitors have been developed, they are limited by poor selectivity and nonspecificity. In this review, we focus on the nonspecific actions of PKC inhibitors on cardiovascular ion channels in addition to their PKC-inhibiting functions. The aim of this paper is to urge caution when using PKC inhibitors to block PKC function. This information may help to better understand PKC-related physiological/biochemical studies.

• The Korean Journal of Physiology and Pharmacology
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• v.27 no.4
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• pp.311-323
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• 2023

Ion homeostasis, which is regulated by ion channels, is crucial for intracellular signaling. These channels are involved in diverse signaling pathways, including cell proliferation, migration, and intracellular calcium dynamics. Consequently, ion channel dysfunction can lead to various diseases. In addition, these channels are present in the plasma membrane and intracellular organelles. However, our understanding of the function of intracellular organellar ion channels is limited. Recent advancements in electrophysiological techniques have enabled us to record ion channels within intracellular organelles and thus learn more about their functions. Autophagy is a vital process of intracellular protein degradation that facilitates the breakdown of aged, unnecessary, and harmful proteins into their amino acid residues. Lysosomes, which were previously considered protein-degrading garbage boxes, are now recognized as crucial intracellular sensors that play significant roles in normal signaling and disease pathogenesis. Lysosomes participate in various processes, including digestion, recycling, exocytosis, calcium signaling, nutrient sensing, and wound repair, highlighting the importance of ion channels in these signaling pathways. This review focuses on different lysosomal ion channels, including those associated with diseases, and provides insights into their cellular functions. By summarizing the existing knowledge and literature, this review emphasizes the need for further research in this field. Ultimately, this study aims to provide novel perspectives on the regulation of lysosomal ion channels and the significance of ion-associated signaling in intracellular functions to develop innovative therapeutic targets for rare and lysosomal storage diseases.

• The korean journal of orthodontics
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• v.32 no.3 s.92
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• pp.227-234
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• 2002

In order to investigate the action mechanism of protein kinase C on $K^+$ channel in osteoblastic cell, effects of phorbol 12, 13-dibutyrate on human osteoblast-like cells (G292) were studied by patch clamp technique with cell-attacked configuration. 111 this experiment, 45pS ion channel was dominant in G292 cell line according to their approximate conductances in symmetrical 140mM KCl saline at holding potential of 60mV. In torrent-voltage relationship, reversal potential was 5.5mV at the condition of potassium enriched saline in the pipette and -27 mV at the condition of standard extracellular saline In the pipette. Phorbol 12, 13-dibutyrate 10nM increased the open probability of 45pS channel and staurosporine, an inhibitor of protein kinase C, suppressed this effect. Phorbol 12,13-dibutyrate moved the reversal potential of 45pS channel to more negative potential and increased the single channel current at the same membrame potential. In order to check the activation of protein kinase C in G292 cell by phorbol 12,13-dibutyrate, western blot of protein kinase C was performed. Phorbol 12,13-dibutyrate $0.1{\mu}M$ translocated protein kinase C from cellular compartment to membrane compartment of the cell. These findings suggest that phorbol 12,13-dibutyrate, one of phorbol esters, activate 45pS channel In G292 cell and affect cell membrane potential, that regulate cellular function.