• 한국생물물리학회:학술대회논문집
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• 한국생물물리학회 1999년도 학술발표회 진행표 및 논문초록
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• pp.43-43
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• 1999

Signal peptides of secretary proteins interact with various membranes and non-membrane components during the translocation. We investigated the interaction of signal peptides of ribose binding protein (RBP) with Escherichia coli (E.coli) signal recognition particle (SRP), SecA and lipid bilayer. Previous studies showed that the functional signal peptides inhibit the GTPase activity of E.coli SRP which consisted of F로 and 4.5S RNA.(omitted)

• 한국생물물리학회:학술대회논문집
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• 한국생물물리학회 1999년도 학술발표회 진행표 및 논문초록
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• pp.56-56
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• 1999

Change of membrane property can affect the activity of membrane proteins. In this work, we investigated the single channel properties of large conductance $Ca^{2+}$-activated $K^{+}$(BK) channels in planar lipid bilayers of different thickness. First, we recorded the activity of single BK channels from rat skeletal muscle incorporated into the control bilayer, then increased the bilayer thickness by perfusing the recording solution with the one saturated with n-pentane, or reduced the thickness by adding diheptanoylphosphatidylcholine (di$C_{7:0}$PC) to the recording soluton.(omitted)

• Journal of Photoscience
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• 제12권2호
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• pp.67-73
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• 2005

Fluorescent probe techniques were used to evaluate the effect of dibucaine.HCl on the physical properties (transbilayer asymmetric lateral mobility, annular lipid fluidity and protein distribution) of synaptosomal plasma membrane vesicles (SPMV) isolated from bovine cerebral cortex. An experimental procedure was used based on selective quenching of 1,3-di(l-pyrenyl)propane (Py-3-Py) by trinitrophenyl groups, and radiationless energy transfer from the tryptophans of membrane proteins to Py-3-Py. Dibucaine.HCl increased the bulk lateral mobility, and annular lipid fluidity in SPMV lipid bilayers, and had a greater fluidizing effect on the inner monolayer than the outer monolayer. The magnitude of increasing effect on annular lipid fluidity in SPMV lipid bilayer induced by dibucaine.HCl was significantly far greater than magnitude of increasing effect of the drug on the lateral mobility of bulk SPMV lipid bilayer. It also caused membrane proteins to cluster. These effects of dibucaine.HCl on neuronal membranes may be responsible for some, though not all, of the local anesthetic actions of dibucaine.HCl.

• 대한수의학회지
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• 제32권4호
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• pp.543-553
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• 1992

Properties of unitary ATP-sensitive $K^+$ channels were studied using planar lipid bilayer technique. Vesicles were prepared from bullfrog (Rana catesbeiana) skeletal muscle. ATP-sensitive $K^+$ (K (ATP)) channels were identified by their unitary conductance and sensitivity to ATP. In the symmetrical solution containing 200mM KCI, 10mM Hepes, 1mM EGTA and pH 7.2, single K (ATP) channels showed a linear current-voltage relations with slight inward rectification. Slope conductance at reversal potential was $60.1{\pm}0.43$ pS(n=3)). Micromolar ATP reversibly inhibited the channel activity when applied to the cytoplasmic side. In the range of -50~＋50 mV, the channel activity was not voltage-dependent, but the channel gating within a burst was more frequent at negative voltage range. Varying the concentrations of external/internal KCl(mM) to 40/200, 200/200, 200/100 and 200/40 shifted reversal potentials to $-30.8{\pm}2.9$(n=3), $-1.1{\pm}2.7$(n=3), 10.5 and 30.6(mV), respecrivety. These reversal potentials were close to the expected values by the Nernst equation, indicating nearly ideal selectivity for $K^+$ over $Cl^-$. Under bi-ionic conditions of 200mM external test ions and 200mM internal $K^+$, the reversal potentials for each test ion/K pair were measured. The measured reversal potentials were used for the calculation of the releative permeability of alkali cations to $K^+$ ions using the Goldman-Hodgkin-Katz equation. The permeability sequence of 5 cations relative to $K^+$ was $K^+$(1), $Rb^+$(0.49), $Cs^+$(0.27), $Na^+$(0.027) and $Li^+$(0.021). This sequence was recognized as Eisenman's selectivity sequence IV. In addition, modelling the permeation of $K^+$ ion through ATP-sensitive $K^+$ channel revealed that a 3-barrier 2-site multiple occupancy model can reasonably predict the observed current-voltage relations.

• BMB Reports
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• 제37권4호
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• pp.460-465
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• 2004

A 16-residue polypeptide model with the sequence acetyl-YALSLAATLLKEAASL-OH was derived by rational de novo peptide design. The designed sequence consists of amino acid residues with high propensity to adopt an alpha helical conformation, and sequential order was arranged to produce an amphipathic surface. The designed sequence was chemically synthesized using a solid-phase method and the polypeptide was purified by reverse-phase liquid chromatography. Molecular mass analysis by electro-spray ionization mass spectroscopy confirmed the correct designed sequence. Structural characterization by circular dichroism spectroscopy demonstrated that the peptide adopts the expected alpha helical conformation in 50% acetonitrile solution. Liposome binding assay using Small Unilamellar Vesicle (SUV) showed a marked release of entrapped glucose by interaction between the lipid membrane and the tested peptide. The channel-forming activity of the peptide was revealed by a planar lipid bilayer experiment. An analysis of the conducting current at various applied potentials suggested that the peptide forms a cationic ion channel with an intrinsic conductance of 188 pS. These results demonstrate that a simple rational de novo design can be successfully employed to create short peptides with desired structures and functions.

• 분석과학
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• 제26권6호
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• pp.395-400
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• 2013

얇은 유리판 위에서 자동적으로 정렬되는 인지질 분자의 정렬도에 대한 수분 및 항균성 펩타이드의 효과를 고체 핵자기 공명 분광법을 이용하여 조사하였다. 순수한 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphotidylcholine (POPC) 인지질 만을 유리판 위에서 정렬시킬 때에는 직접적인 물의 투입 없이 95% 상대 습도에서 수일간 수화시키는 것만으로 충분했다. 하지만, protegrin-1 (PG-1)과 같은 항균성 펩타이드가 혼합되어 있는 인지질의 경우에는 95%의 상대 습도에서 수일간 수화시키는 것과 약간의 물을 시료에 직접 투입하여 수화시키는 것이 크게 달라진다는 것을 확인하였다. 충분한 양의 물이 투입되었을 때, 지질 이중막 위에서 인지질 분자들의 표면 움직임이 매우 활발하였다. 순수한 POPC 분자들이 얇은 유리판 위에서 정렬되는 시간에 비해 항균성 펩타이드가 혼합되어 있는 경우에는 POPC 분자들이 평형상을 이루어질 때까지는 상당한 시간이 필요함을 확인하였다.

• The Korean Journal of Physiology
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• 제22권2호
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• pp.163-177
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• 1988

세포막을 구성하고 있는 지질의 물리학적 성상이 단백질의 세포막 속으로의 삽입과정 및 단백질의 기능에 미치는 영향을 관찰하기 위하여 골격근의 근세망(SR)으로부터 $Ca^{2+}-ATPase$ 단백질을 분리한 후 이를 세포막의 주 구성성분인 포스파티딜콜린(PC)과 포스파티딜에타노라민(PE)의 혼합지질과 재조합(reconstitution)시켰다. 이와같이 인공적으로 재조합된 구조물에서 $Ca^{2+}-ATPase$의 기능을 측정하기 위하여 칼슘지시색소인 아르세나죠III(AIII)를 이용한 분광방법과 방사선동위원소를 이용한 여과법으로 칼슘흡수율을 측정하였고 또한 ATP 가수분해 능력을 측정하였다. 실험결과 칼슘의 흡수율은 포스파티딜코린의 함량이 많은 혼합지질과 재조합시킬 때에 증가하였고, ATP 가수분해 능력은 포스파티딜함량이 25%까지는 포스타피딜코린의 양에 비례하여 증가하였으나 50%이상에서는 약간 감소하는 경향을 보였다. 한편 지질세포막속으로 단백질이 삽입되는 양은 포스파티딜 함량이 25%일 때 최고의 값을 보였으며 함량이 그 이하 또는 이상일 때는 감소하였다. 이상의 실험결과로보아 단백질의 기능은 세포막이 "bilayer" 구조를 갖출때에 증가하고 세포막속으로 단백질이 삽입되는 양은 세포막이 "non-bilayer" 구조를 형성할 때에 증가함을 알 수 있다.

• BMB Reports
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• 제37권5호
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• pp.603-611
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• 2004

Fluorescence probes located in different membrane regions were used to evaluate the effects of chlorpromazine HCl on structural parameters (transbilayer lateral mobility, annular lipid fluidity, protein distribution, and lipid bilayer thickness) of synaptosomal plasma membrane vesicles (SPMVs) isolated from bovine cerebral cortex. The experimental procedure was based on the selective quenching of 1,3-di(1-pyrenyl)propane (Py-3-Py) by trinitrophenyl groups, radiationless energy transfer from the tryptophan of membrane proteins to Py-3-Py, and energy transfer from Py-3-Py monomers to 1-anilinonaphthalene-8-sulfonic acid (ANS). In this study, chlorpromazine HCl decreased the lateral mobility of Py-3-Py in a concentration dependent-manner, showed a greater ordering effect on the inner monolayer than on the outer monolayer, decreased annular lipid fluidity in a dose dependent-manner, and contracted the membrane lipid bilayer. Furthermore, the drug was found to have a clustering effect on membrane proteins.

• The Korean Journal of Physiology and Pharmacology
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• 제13권1호
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• pp.39-47
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• 2009

Gaegurin 4(GGN 4), an antimicrobial peptide isolated from a Korean frog, is five times more potent against Gram-positive than Gram-negative bacteria, but has little hemolytic activity. To understand the mechanism of such cell selectivity, we examined GGN4-induced $K^+$ efflux from target cells, and membrane conductances in planar lipid bilayers. The $K^+$ efflux from Gram-positive M. luteus(2.5 ${\mu}g/ml$) was faster and larger than that from Gram-negative E. coli(75 ${\mu}g/ml$), while that from RBC was negligible even at higher concentration(100 ${\mu}g/ml$). GGN4 induced larger conductances in the planar bilayers which were formed with lipids extracted from Gram-positive B. subtilis than in those from E. coli(p<0.01), however, the effects of GGN4 were not selective in the bilayers formed with lipids from E. coli and red blood cells. Addition of an acidic phospholipid, phosphatidylserine to planar bilayers increased the GGN4-induced membrane conductance(p<0.05), but addition of phosphatidylcholine or cholesterol reduced it(p<0.05). Transmission electron microscopy revealed that GGN4 induced pore-like damages in M. luteus and dis-layering damages on the outer wall of E. coli. Taken together, the present results indicate that the selectivity of GGN4 toward Gram-positive over Gram-negative bacteria is due to negative surface charges, and interaction of GGN4 with outer walls. The selectivity toward bacteria over RBC is due to the presence of phosphatidylcholine and cholesterol, and the trans-bilayer lipid asymmetry in RBC. The results suggest that design of selective antimicrobial peptides should be based on the composition and topology of membrane lipids in the target cells.

• The Korean Journal of Physiology and Pharmacology
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• 제2권4호
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• pp.529-539
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• 1998

The effects of membrane surface charge originated from lipid head groups on ion channels were tested by analyzing the activity of single large conductance $Ca^{2+}-activated\;K^+$ (maxi K) channel from rat skeletal muscle. The conductances and open-state probability ($P_o$) of single maxi K channels were compared in three types of planar lipid bilayers formed from a neutral phosphatidylethanolamine (PE) or two negatively-charged phospholipids, phosphatidylserine (PS) and phosphatidylinositol (PI). Under symmetrical KCl concentrations $(3{\sim}1,000\;mM)$, single channel conductances of maxi K channels in charged membranes were $1.1{\sim}1.7$ times larger than those in PE membranes, and the differences were more pronounced at the lower ionic strength. The average slope conductances at 100 mM KCl were $251{\pm}9.9$, $360{\pm}8.7$ and $356{\pm}12.4$ $(mean{\pm}SEM)$ pS in PE, PS and PI membranes respectively. The potentials at which $P_o$ was 1/2, appeared to have shifted left by 40 mV along voltage axis in the membranes formed with PS or PI. Such shift was consistently seen at pCa 5, 4.5, 4 and 3.5. Estimation of the effect of surface charge from these data indicated that maxi K channels sensed the surface potentials at a distance of $8{\sim}9\;{\AA}$ from the membrane surface. In addition, similar insulation distance ($7{\sim}9\;{\AA}$) of channel mouth from the bilayer surface charge was predicted by a 3-barrier-2-site model of energy profile for the permeation of $K^+$ ions. In conclusion, despite the differences in structure and fluidity of phospholipids in bilayers, the activities of maxi K channels in two charged membranes composed of PS or PI were strikingly similar and larger than those in bilayers of PE. These results suggest that the enhancement of conductance and $P_o$ of maxi channels is mostly due to negative charges in the phospholipid head groups.