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Y K Shin

Publications and source records attributed to Y K Shin.

At least 91 records · Page 5Linked to original sources
PubMed1995

Effect of dantrolene sodium on contractility of isolated human uterine muscle.

The administration of intravenous dantrolene in a parturient susceptible to malignant hyperthermia has been associated with post partum uterine atony. We examined the effect of dantrolene sodium for injection (Dantrium Intravenous) on spontaneous contractility of uterine smooth muscle from women in term pregnancy in an isolated preparation. Dantrolene sodium for injection at 5 microg/ml and 10 microg/ml had no effect on the spontaneous contractility of the uterine muscle preparations. At a cumulative concentration of 20 microg/ml, a mild depression (16 +/- 14%) in the frequency of spontaneous contractions was noted. However, a similar depression in the muscle preparations treated with mannitol suggests that the depression observed with the dantrolene was likely due to the mannitol that was included in the dantrolene formulation rather than to dantrolene sodium itself. We conclude that dantrolene sodium has no effect on the spontaneous contractility of uterine smooth muscle. The depression of uterine muscle activity observed with dantrolene for injection appears attributable to the mannitol.

Journal Article↗
PubMed1995

High-level expression from a cytomegalovirus promoter in macrophage cells.

To identify vectors that provide optimal gene expression in human hematopoietic cells, we investigated retroviruses containing the adenosine deaminase (ADA) gene under the transcriptional control of the promoters/enhancers of Moloney murine leukemia virus and the human cytomegalovirus (CMV) immediate-early gene. ADA expression was monitored in transduced human multipotential promyelocytic leukemic HL-60 cells and human monocytic leukemic THP-1 cells. HL-60 cells can be induced by phorbol ester to differentiate into macrophage lineage cells and by retinoic acid into granulocytic cells. THP-1 cells undergo phorbol ester-induced differentiation to macrophage cells. In LNCA-transduced HL-60 derived macrophage cells, ADA controlled by the CMV promoter was expressed at 100.0 mumol/hr.mg, in contrast to 1.2 mumol/hr.mg from LN-transduced control cells. LNCA-transduced THP-1 macrophage cells showed a similar increase in ADA activity over control cells. These elevated enzyme activities were confirmed by Northern blots, which showed substantial increases in ADA mRNA derived from the CMV promoter. This suggests use of the CMV promoter for gene therapy targeted at macrophages, as, for example, in the treatment of lysosomal storage disorders such as Gaucher disease. These inducible cell lines have allowed the evaluation of transduced gene expression in proliferating and differentiating hematopoietic cells that may serve as an in vitro model of bone marrow-targeted gene therapy.

Adenosine Deaminase↗
PubMed1994

Topology of an amphiphilic mitochondrial signal sequence in the membrane-inserted state: a spin labeling study.

To investigate the interaction of the presequence of the precursor of yeast cytochrome C oxidase subunit IV (COX IV) with phospholipid membranes, a series of single- and double-cysteine-substituted peptide variants derived from the 25-residue NH2-terminal presequence has been synthesized and modified with nitroxide spin labels. The immersion depth, orientation, and secondary structure of the peptide in the POPC bilayer containing 10 mol % POPG were determined using electron paramagnetic resonance (EPR) spectroscopy. EPR saturation analysis of singly labeled variants reveals that the nitroxides attached to the NH2-terminal region of the peptide insert into the acyl chain region of the bilayer, approximately 13 A deep from the membrane surface. EPR line shape analysis of doubly labeled variants indicates that the peptide predominantly exists as an extended conformation, with little secondary structure. The experimental results, together with the energetic consideration of peptide-bilayer interactions, suggest that the presequence is located near the interface between the head group region and the acyl chain region, such that the hydrophobic side chains are solvated by the acyl chains and the charged side chains extended toward the polar environment at the bilayer surface.

Amino Acid Sequence↗
PubMed1994

Insertion of a coiled-coil peptide from influenza virus hemagglutinin into membranes.

The trimeric protein hemagglutinin (HA) of the influenza viral envelope is essential for cell entry. To investigate the interaction of HA with membranes, two 40-residue, cysteine-substituted peptides comprising the loop region and the first part of the coiled-coil stem were synthesized and modified with a nitroxide spin label. Electron paramagnetic resonance analysis revealed that the peptide inserts reversibly into phospholipid vesicles under endosomal pH conditions. This result suggests that some or all of the long coiled-coil trimer of HA may insert into membranes, which could bring the viral and cell membranes closer together and facilitate fusion.

Amino Acid Sequence↗
PubMed1993

Colicin E1 binding to membranes: time-resolved studies of spin-labeled mutants.

To investigate the mechanism of interaction of the toxin colicin E1 with membranes, three cysteine substitution mutants and the wild type of the channel-forming fragment were spin labeled at the unique thiol. Time-resolved interaction of these labeled proteins with phospholipid vesicles was investigated with stopped-flow electron paramagnetic resonance spectroscopy. The fragment interacts with neutral bilayers at low pH, indicating that the interaction is hydrophobic rather than electrostatic. The interaction occurs in at least two distinct steps: (i) rapid adsorption to the surface; and (ii) slow, rate-limiting insertion of the hydrophobic central helices into the membrane interior.

Adsorption↗
PubMed1993

Thermodynamics and dynamics of phosphatidylcholine-cholesterol mixed model membranes in the liquid crystalline state: effects of water.

A method for obtaining the thermodynamic activity of each membrane component in phosphatidylcholine (PC)/cholesterol mixtures, that is based upon ESR spin labeling is examined. The thermodynamic activity coefficients, gamma PC and gamma chol, for the PC and cholesterol, respectively, are obtained from the measured orientational order parameters, SPC and S(chol), as a function of cholesterol content for a spin-labeled PC and the sterol-type cholestane spin probe (CSL), respectively, and the effects of water concentration are also considered. At water content of 24 weight%, the thermodynamics of DMPC/cholesterol/water mixtures in the liquid-crystalline state may be treated as a two-component solution ignoring the water, but at lower water content the role of water is important, especially at lower cholesterol concentrations. At lower water content (17 wt%), gamma chol decreases with increasing cholesterol content which implies aggregation. However, at higher water content (24 wt%), gamma chol is found initially to increase as a function of cholesterol content before decreasing at higher cholesterol content. This implies a favorable accommodation for the cholesterol in the membrane at high water and low cholesterol content. Good thermodynamic consistency according to the Gibbs-Duhem equation was obtained for gamma PC and gamma chol at 24 wt% water. The availability of gamma chol (and gamma PC) as a function of cholesterol concentration permits the estimate of the boundary for phase separation. The rotational diffusion coefficients of the labeled PC and of CSL were also obtained from the ESR spectra. A previously proposed universal relation for the perpendicular component of the rotational diffusion tensor, R perpendicular, for CSL in PC/cholesterol mixtures (i.e., R perpendicular = R0 perpendicular exp(-AS2chol/RT)) is confirmed. A change in composition of cholesterol or of water for DMPC/cholesterol/water mixtures affects R perpendicular only through the dependence of S(chol) on the composition. In particular, the amount of water affects the membrane fluidity, monitored by R perpendicular for CSL, solely by the structural changes it induces in the membrane for the compositions studied. Rotational diffusion for the labeled PC is found to be more complex, most likely due to the combined action of the internal modes of motion of the flexible chain and of the overall molecular reorientation.

Biophysical Phenomena↗
PubMed1993

Molecular systematics of the genus Zoogloea and emendation of the genus.

Phylogenetic relationships among strains of Zoogloea and related taxa were determined by 16S rDNA sequencing and genomic DNA hybridization techniques. The 16S rRNA gene was amplified by the polymerase chain reaction with a pair of eubacterial consensus primers and sequenced directly by using an automated fluorescent DNA sequencer. Sequence comparisons and distance matrix tree analysis revealed that Zoogloea ramigera IAM 12136 (= N. C. Dondero 106, type strain) and Zoogloea sp. ATCC 19324 formed a lineage with Rhodocyclus purpureus in the beta subclass of Proteobacteria. Z. ramigera IAM 12670 (= P. R. Dugan 115) was shown to belong to another cluster with Alcaligenes eutrophus and Pseudomonas cepacia in the beta subclass. In contrast, Z. ramigera IAM 12669 (= K. Crabtree I-16-M) proved to be a member of the alpha subclass of the Proteobacteria, closely related to Agrobacterium tumefaciens. Genomic DNA hybridization studies also showed that there is genetic diversity among the strains currently designated Z. ramigera, but typical Zoogloea strains, characterized by their production of rhodoquinones, are highly related to each other and can be regarded as a single species. On the basis of the molecular data, together with the early phenotypic and chemotaxonomic information, we have emended the generic description of Zoogloea.

Base Composition↗
PubMed1992

Isoprenoid quinones and fatty acids of Zoogloea.

Nine Zoogloea strains including the type strain of Z. ramigera (IAM 12136 = ATCC 19544 = N.C. Dondero 106) and newly isolated strains were investigated for isoprenoid quinone composition and whole-cell fatty acid profiles. Seven of the tested strains, having phenotypic properties typical of Zoogloea, were characterized by their production of both ubiquinone-8 and rhodoquinone-8 as major quinones, whereas the remaining two strains, Z. ramigera IAM 12669 (= K. Crabtree I-16-M) and IAM 12670 (= P.R. Dugan 115), formed ubiquinone-10 and ubiquinone-8, respectively, as the sole quinone. All rhodoquinone-producing strains contained palmitoleic acid and 3-hydroxy-decanoic acid as the major components of nonpolar and hydroxylated fatty acids, respectively. Marked differences were noted in the fatty acid composition between the strains with and without rhodoquinones. The chemotaxonomic data suggested that the rhodoquinone-lacking strains should be excluded from the genus Zoogloea. Since there have been no reliable taxonomic tools for Zoogloea, rhodoquinone analysis may provide a new criterion of great promise for identifying Zoogloea strains.

Chromatography, High Pressure Liquid↗
PubMed1992

Determination of electrostatic potentials at biological interfaces using electron-electron double resonance.

A new general method for the determination of electrostatic potentials at biological surfaces is presented. The approach is based on measurement of the collision frequency of a charged nitroxide in solution with a nitroxide fixed to the surface at the point of interest. The collision frequency is determined with 14N:15N double label electron-electron double resonance (ELDOR). As a test, the method is shown to give values for phospholipid bilayer surface potentials consistent with the Gouy-Chapman theory, a simple model shown by many independent tests to accurately describe charged, planar surfaces. In addition, the method is applied to determine the electrostatic potential near the surface of DNA. The results indicate that the potential is significantly smaller than that predicted from Poisson-Boltzmann analysis, but is in qualitative agreement with that predicted by Manning's theory of counter ion condensation. The method is readily extended to measurement of surface potentials of proteins.

Aminacrine↗
PubMed1992

Postpartum transient blindness probably related to undiagnosed brain tumor.

Nausea and vomiting are so frequent under regional anesthesia during parturition that these symptoms can be misleading and may cause the clinician to overlook a potential intracranial lesion. In the case we have reported, it was not until the onset of blindness that an intracranial lesion was suspected. Although the incidence of brain tumor during pregnancy is low, it does occur with various manifestations and has been associated with blindness. Therefore, prompt neurologic investigation may be warranted in patients complaining of visual changes, even though transient blindness can occur in the absence of a neurologic lesion.

Adult↗
PubMed1991

Microscopic versus macroscopic diffusion in model membranes by electron spin resonance spectral-spatial imaging.

The macroscopic and the microscopic diffusion coefficients of a phospholipid spin label (16-PC) in the model membrane 1-palmitoyl-2-oleoyl-sn-glycero-phosphatidylcholine have been measured simultaneously in the same sample utilizing the new technique of spectral-spatial electron spin resonance imaging. The macroscopic diffusion coefficient Dmacro for self-diffusion of 16-PC spin label is obtained from imaging the concentration profiles as a function of time, and it is (2.3 +/- 0.4) x 10(-8) cm2/s at 22 degrees C. The microscopic diffusion coefficient Dmicro for relative diffusion of the spin probes is obtained from the variation of the spectral line broadening with spin label concentration, which is due to spin-spin interactions. Dmicro is found to be substantially greater than Dmacro for the same sample at the same conditions, and is estimated to be at least (1.0 +/- 0.4) x 10(-7) cm2/s. Possible sources for their difference are briefly discussed in terms of the models used for Dmicro.

Cyclic N-Oxides↗
PubMed1990

Recombinant RNA polymerase: inducible overexpression, purification and assembly of Escherichia coli rpo gene products.

The genes, rpoA, rpoB and rpoC of Escherichia coli, which encode the RNA polymerase alpha-, beta- and beta'-subunits, respectively, have been individually placed on expression plasmids under the control of the bacteriophage T7 promoter. Induction of the T7 RNA polymerase gene in host cells harboring each of the three plasmids resulted in the extensive overproduction of the three polypeptides. The overproduced subunits were purified and assembled into a functional enzyme, whose specific activity and dependence on the sigma-factor were indistinguishable from native RNA polymerase purified by conventional methods.

Cloning, Molecular↗
PubMed1990

Dynamics of phosphatidylcholine-cholesterol mixed model membranes in the liquid crystalline state.

The effects of cholesterol on the dynamics of cholestane spin probe (CSL) in various phosphatidylcholine-cholesterol mixed model membranes are examined. The lateral diffusion, D of CSL in DMPC/POPC/cholesterol ternary mixtures, is measured utilizing an improved dynamic imaging electron spin resonance method. It shows a factor of two decrease at 10 mol % and 25 degrees C, whereas it shows only a 40% decrease at 50 mol % and 50 degrees C. A comparison with results in POPC/cholesterol mixtures, which show a stronger effect of cholesterol on D, indicates that acyl chain unsaturation leads to stronger self association of cholesterol in PC model membranes. An S2CSL dependence of the activation energy for D, has been confirmed for the DMPC/POPC/cholesterol mixtures. Here SCSL is the order parameter for CSL. A similar correlation of R perpendicular, the perpendicular component of the rotational diffusion coefficient, with SCSL, which is true for all three mixtures (DMPC/cholesterol, POPC/cholesterol, and DMPC/POPC/cholesterol) we have studied, is also found. These are associated with the effects of enhanced local ordering on the free volume needed for translation and reorientation. Such correlations of dynamic properties D and R perpendicular with the thermodynamic quantity S, as well as the consistent interpretations of the effect of acyl chain unsaturation on the dynamics in terms of the activity coefficients, strongly emphasize the interrelation between the dynamic structure and the thermodynamics of the PC/cholesterol mixtures.

Cholesterol↗
PubMed1989

Thermodynamics of phosphatidylcholine-cholesterol mixed model membranes in the liquid crystalline state studied by the orientational order parameter.

It is shown that good estimates of the activity of cholesterol in phosphatidylcholine-cholesterol mixed model membranes are obtained by examining the orientational order parameter S of cholestane spin probe (CSL) that is obtained from electron spin resonance by spectral simulation. By introducing thermodynamic stability conditions of liquid mixtures, the variation of activity (or S) as a function of cholesterol mole fraction is utilized to predict the concentration at which the phase separation occurs. These results for DMPC and cholesterol binary mixtures agree very well with those of Tempo-partitioning experiments. The comparison of activity coefficients and the phase boundary in DMPC/cholesterol mixtures with those of POPC/cholesterol mixtures suggests that acyl chain unsaturation leads to poorer mixing of cholesterol in phosphatidylcholine model membranes at higher temperatures (i.e., greater than 35 degrees C). In ternary solutions of DMPC, POPC, and cholesterol, it is found that cholesterol shows less deviation from ideality than in either of the two binary mixtures, and this implies that the phase separation occurs at higher cholesterol concentration than in either of the two binary mixtures. The present analysis suggests that there may not be a critical point in DMPC/cholesterol mixtures, even though phase separation does occur.

Cholesterol↗
PubMed1989

Dynamic imaging of lateral diffusion by electron spin resonance and study of rotational dynamics in model membranes. Effect of cholesterol.

The effects of cholesterol on the dynamics and the structural properties of two different spin probes, the sterol type CSL and the phospholipid type 16-PC, in POPC/cholesterol oriented multilayer model membranes were examined. Our results are consistent with a nonideal solution containing cholesterol-rich clusters created by the self association of cholesterol in POPC model membranes. The lateral diffusion coefficient D of the spin probes was measured over the temperature range of 15 to 60 degrees C and over the concentration range of 0 to 30 mol% of cholesterol in the model membrane by the electron spin resonance (ESR) imaging method. The rotational diffusion coefficients (including R perpendicular) and the order parameter S were determined utilizing a nonlinear least square ESR spectral simulation method. D, R perpendicular and S of CSL deviate considerably from linear dependence on mole percent cholesterol. The D of CSL was decreased by a factor of four at 15 degrees C and a factor of two at 60 degrees C for concentrations of cholesterol over 10 mol %, whereas those of 16-PC were hardly affected. Cholesterol decreased R perpendicular by a factor of 10 at 30 mol % of cholesterol, but it increased slightly that of 16-PC. A significant increase of S for CSL due to the presence of cholesterol was observed. It is shown how the difference in variation of S for CSL vs. 16-PC with composition may be interpreted in terms of their respective activity coefficients, and how a single universal linear relation is obtained for the S of both probes in terms of a scaled temperature. Simple but general correlations of D and of R perpendicular with S were also found, which aid in the interpretation of these diffusion coefficients.

Cholesterol↗