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Biomedical subjects

J S Yoo

Publications and source records attributed to J S Yoo.

At least 37 records · Page 2Linked to original sources

Identification of acylated glycoglycerolipids from a cyanobacterium, Synechocystis sp., by tandem mass spectrometry.

Acylated glycoglycerolipids were identified in the total lipid extract from cyanobacterium Synechocystis sp. PCC 6803. These compounds have a palmitoyl group esterified to the hydroxyl group at the C-6 position of the terminal glycosyl moiety of digalactosyl monoacylglycerol and digalactosyl diacylglycerol. Their structural elucidation was accomplished by tandem mass spectrometry coupled with fast atom bombardment ionization. Acylated digalactosyl monoacylglycerol has a structure of 1-hydroxy-2-palmitoyl-3-O-[(6-O-palmitoyl)-alpha-D-galactopyranosyl -(1-->6)-beta-D-galactopyranosyl]-sn-glycerol. This compound has not been reported previously.

Acylation↗

Vesicle-associated membrane protein 4 is implicated in trans-Golgi network vesicle trafficking.

The trans-Golgi network (TGN) plays a pivotal role in directing proteins in the secretory pathway to the appropriate cellular destination. VAMP4, a recently discovered member of the vesicle-associated membrane protein (VAMP) family of trafficking proteins, has been suggested to play a role in mediating TGN trafficking. To better understand the function of VAMP4, we examined its precise subcellular distribution. Indirect immunofluorescence and electron microscopy revealed that the majority of VAMP4 localized to tubular and vesicular membranes of the TGN, which were in part coated with clathrin. In these compartments, VAMP4 was found to colocalize with the putative TGN-trafficking protein syntaxin 6. Additional labeling was also present on clathrin-coated and noncoated vesicles, on endosomes and the medial and trans side of the Golgi complex, as well as on immature secretory granules in PC12 cells. Immunoprecipitation of VAMP4 from rat brain detergent extracts revealed that VAMP4 exists in a complex containing syntaxin 6. Converging lines of evidence implicate a role for VAMP4 in TGN-to-endosome transport.

Animals↗

Three novel proteins of the syntaxin/SNAP-25 family.

Intracellular membrane traffic is thought to be regulated in part by soluble N-ethylmaleimide-sensitive factor-attachment protein receptors (SNAREs) through the formation of complexes between these proteins present on vesicle and target membranes. All known SNARE-mediated fusion events involve members of the syntaxin and vesicle-associated membrane protein families. The diversity of mammalian membrane compartments predicts the existence of a large number of different syntaxin and vesicle-associated membrane protein genes. To further investigate the spectrum of SNAREs and their roles in membrane trafficking we characterized three novel members of the syntaxin and SNAP-25 (synaptosome-associated protein of 25 kDa) subfamilies. The proteins are broadly expressed, suggesting a general role in vesicle trafficking, and localize to distinct membrane compartments. Syntaxin 8 co-localizes with markers of the endoplasmic reticulum. Syntaxin 17, a divergent member of the syntaxin family, partially overlaps with endoplasmic reticulum markers, and SNAP-29 is broadly localized on multiple membranes. SNAP-29 does not contain a predicted membrane anchor characteristic of other SNAREs. In vitro studies established that SNAP-29 is capable of binding to a broad range of syntaxins.

Amino Acid Sequence↗

Seven novel mammalian SNARE proteins localize to distinct membrane compartments.

Soluble N-ethylmaleimide-sensitive factor-attachment protein receptor (SNARE) proteins of the vesicle-associated membrane protein (VAMP) and syntaxin families play a central role in vesicular trafficking through the formation of complexes between proteins present on vesicle and target membranes. Formation of these complexes is proposed to mediate aspects of the specificity of vesicle trafficking and to promote fusion of the lipid bilayers. In order to further understand the molecular mechanisms that organize membrane compartments, we have characterized seven new mammalian proteins of the VAMP and syntaxin families. The proteins are broadly expressed; however, syntaxin 13 is enriched in brain and VAMP 8 in kidney. The seven novel SNAREs localize in distinct patterns overlapping with Golgi, endosomal, or lysosomal markers. Our studies support the hypothesis that evolutionary radiation of these two gene families gave rise to sets of proteins whose differential expression and combinatorial associations define and organize the membrane compartments of cells.

Amino Acid Sequence↗

Malononitrile as a new derivatizing reagent for high-sensitivity analysis of oligosaccharides by electrospray ionization mass spectrometry.

A new method for the high-sensitivity analysis of oligosaccharides by negative ion electrospray ionization mass spectrometry was developed through a chemical derivatization of oligosaccharides. Oligosaccharides were derivatized to dinitrile compounds from the reaction with malononitrile under mildly basic conditions. The derivative of maltoheptaose was detected mainly as the [M-2H]2- ion in negative ion mode with 20 fmol sensitivity, even in unpurified samples. In this malononitrile derivatization method, no inorganic reagent, other than sodium hydroxide as a base catalyst, is used. Also, because excess ligand (malononitrile) is volatile, high sensitivity detection is realized without any solvent extraction or chromatographic purification. The detection limit can also be decreased by simple on-line cartridge filtration to 200 attomol which is 10(5) times better than that of free maltoheptaose. Structural information for oligosaccharide derivatives was obtained by collision induced dissociation. This malononitrile derivatization method is convenient and efficient for the sensitive analysis of oligosaccharides.

Carbohydrate Sequence↗

Subunit structure of the mammalian exocyst complex.

The exocyst is a protein complex required for the late stages of secretion in yeast. Unlike the SNAREs (SNAP receptors), important secretory proteins that are broadly distributed on the target membrane, the exocyst is specifically located at sites of vesicle fusion. We have isolated cDNAs encoding the rexo70, rsec5, and rsec15 subunits of the mammalian complex. The amino acid sequences encoded by these genes are between 21% and 24% identical to their yeast homologs. All three genes are broadly expressed and multiple transcripts are observed for rexo70 and rsec15. Characterization of cDNAs encoding the 84-kDa subunit of the mammalian complex revealed a novel protein. mAbs were generated to the mammalian rsec6 subunit of the exocyst complex. rsec6 immunoreactivity is found in a punctate distribution at terminals of PC12 cell processes at or near sites of granule exocytosis.

Amino Acid Sequence↗

Modulation of the catalytic activity of brain succinic semialdehyde reductase by reaction with pyridoxal 5'-phosphate.

An NADPH-dependent succinic semialdehyde reductase from bovine brain was inactivated by pyridoxal 5'-phosphate. Spectral evidence is presented to indicate that the inactivation proceeds through formation of a Schiff's base with amino groups of the enzyme. After sodium borohydride reduction of the inactivated enzyme, it was observed that 1 mol phosphopyridoxyl residue was incorporated/mol enzyme monomer. The coenzyme, NADPH, protected the enzyme against inactivation by pyridoxal 5'-phosphate. After tryptic digestion of the enzyme modified with pyridoxal 5'-phosphate in the presence and absence of NADPH followed by [1H]NaBH4 reduction, a radioactive peptide absorbing at 310 nm was isolated by reverse-phase HPLC. The amino acid sequence of the peptide identified a portion of the pyridoxal-5'-phosphate-binding site as the region containing the sequence I-L-E-N-I-Q-V-F-X-K, where X indicates that the phenylthiohydantoin amino acid could not be assigned. The missing residue, however, can be designated as a phosphopyridoxyl lysine as interpreted from the result of amino acid composition of the peptide. It is suggested that the catalytic function of succinic semialdehyde reductase is modulated by binding of pyridoxal 5'-phosphate to a specific lysyl residue at or near the coenzyme-binding site of the protein.

Amino Acids↗

MR566A and MR566B, new melanin synthesis inhibitors produced by Trichoderma harzianum. II. Physico-chemical properties and structural elucidation.

New melanin synthesis inhibitors (MR566A and B) and six related known isocyanocyclopentenes were isolated from the fermentation broth of Trichoderma harzianum, and their structures were elucidated by spectroscopic methods. The structures of novel isocyanides, MR566A (1) and B (2), were elucidated as 1-(3-chloro-1,2-dihydroxy-4-isocyano-4-cyclopenten-1-yl)etha nol, 1-(1,2,3-trihydroxy-3-isocyano-4-cyclopenten-1-yl)ethanol, respectively. The structure of novel oxazole, MR93B (9), was elucidated as 4-[(1Z)-3-hydroxy-2-hydroxymethyl-1-propen-1-yl]oxazole.

Cyclopentanes↗

cis-Hydroxyproline inhibits proliferation, collagen synthesis, attachment, and migration of cultured bovine retinal pigment epithelial cells.

PURPOSE: Proliferative vitreoretinopathy (PVR) is characterized by the proliferation and migration of retinal pigment epithelial (RPE) and other cells into the vitreous cavity. The PVR membrane formation also is associated with collagen production by RPE. The authors examined the effect of a proline analog, cis-hydroxyproline (CHP), on proliferation, collagen synthesis, attachment, and migration of bovine RPE in vitro. METHODS: The effect of CHP on cell proliferation was determined as a function of dosage and days in culture by counting the cell numbers on days 3, 6, and 9. Collagen synthesis was determined by trichloroacetic acid precipitation of the radiolabeled samples before and after bacterial collagenase digestion. The attachment assay involved type I collagen or fibronectin substrates or both (2.5 micrograms/well). For migration experiments, RPE cells were removed from a defined area of a confluent culture, and migration was quantitated by counting the number of cells migrating into the denuded area over 30 hours. RESULTS: The addition of CHP inhibited RPE proliferation in both a dose- and a time-dependent manner; collagen synthesis, attachment, and migration also were inhibited by CHP in a dose-dependent manner. When the culture plates were coated with collagen, < 100 micrograms/ml of CHP had no effect on cell attachment. Higher doses of CHP resulted in mild inhibition of attachment on collagen-coated plates. Simultaneous addition of L-proline to the cultures resulted in blockade of these inhibitory effects on proliferation, collagen synthesis, attachment, and migration. CONCLUSIONS: The results show that RPE functions critical to the development of PVR are inhibited by CHP, suggesting the possibility that this drug may have potential clinical application.

Animals↗

Application of fast atom bombardment combined with tandem mass spectrometry to the structural elucidation of O-demethylabierixin and related polyether antibiotics.

O-Demethylabierixin, a new polyether antibiotic, was isolated from a Streptomyces spp. Its structure elucidation was carried out with fast atom bombardment mass spectrometry (FAB-MS) and fast atom bombardment mass spectrometry-mass spectrometry (FAB-MS/MS) by comparing spectral patterns with those of structurally similar polyether compounds, nigericin and abierixin. The collision-induced dissociation (CID) of sodium-adduct and deprotonated molecular ions produced many cyclic ether rings-opened product ions via a series of the dissociative processes. Because of the negative charge of the carboxylate group at the end of the molecule, the charge-remote fragmentation pattern in the negative CID spectra of deprotonated molecular ions was very helpful for complete identification of product ions which are characteristic for cyclic ether structures. From this study, the new polyether antibiotic was identified to be O-demethylabierixin in which the methoxy group of six-membered ether ring in abierixin was replaced by the hydroxy group.

Anti-Bacterial Agents↗

Protective effects of diallyl sulfide on acetaminophen-induced toxicities.

Diallyl sulfide (DAS), a major flavour component of garlic, is known to modulate drug metabolism and may protect animals from chemically induced toxicity and carcinogenesis. In this study the effects of DAS on the oxidative metabolism and hepatotoxicity induced by acetaminophen (APAP) in rats were investigated. In the hepatotoxicity evaluation of Fischer 344 rats there was a dose-dependent increase in the odds of mortality rate by APAP (P = 0.009); DAS treatment significantly protected rats from APAP-related mortality (P = 0.026). Liver toxicity determined by lactate dehydrogenase activity was significantly increased by APAP treatment (0.75 g/kg). Pretreatment with DAS protected animals from APAP-induced liver toxicity in a time- and dose-dependent fashion. Treatment of DAS (50 mg/kg) 3 hr after APAP dosing significantly (P < 0.05) protected rats from APAP-induced liver toxicity. The metabolism of APAP (50 microM) in vitro was significantly inhibited by DAS (0.3-1 mM) in liver microsomes isolated from F344 rats. As the effect of DAS on APAP-induced hepatotoxicity in vivo was observed only when DAS was administered before or shortly after (< 3 hr) APAP dosing, data suggested that the protective effect of DAS is mainly at the metabolic activation step of APAP. However, the possibility that DAS may also have effects on other drug metabolism systems, such as glutathione (GSH) and glutathione S-transferases, cannot be ruled out.

Acetaminophen↗

Hypericin inhibits cell growth and induces apoptosis in retinal pigment epithelial cells: possible involvement of protein kinase C.

Proliferative vitreoretinopathy (PVR) is characterized by the proliferation and migration of retinal pigment epithelial (RPE) cells in the vitreous cavity. The drug hypericin, which is already in clinical use as an antidepressant, has shown promise as an antiviral and antineoplastic agent. To investigate the therapeutic potential of hypericin in PVR, we incubated RPE cells in standard medium with various serum concentrations containing 0.5 to 5 microM hypericin. In some experiments we studied the effects of hypericin in conjunction with the RPE growth stimulating cytokine tumor necrosis factor alpha (TNF-alpha). Dose-dependent inhibition of RPE cell proliferation with IC50 values of 0.7 microM and 3.3 microM in 1% and 5% serum respectively, was found. Even in conjunction with TNF-alpha, hypericin inhibited RPE proliferation with an IC50 value of 1.5 microM. The drug inhibited PKC activity in cells treated with a 2.5 microM dose by 72% after 30 min and by 100% after 180 min. Finally, hypericin induced RPE cells to undergo apoptotic cell death, as shown by the presence of DNA laddering. These results suggest that hypericin may have potential as a therapeutic drug for PVR and that its antiproliferative and apoptotic effects on RPE cells in vitro are in part mediated by PKC.

Animals↗

Octoxynol presence in bear-bile.

The presence of octoxynol from dried bear-bile was examined. Octoxynol was coextracted when glycolipids by Folch-Suzuki partition method. Octoxynol formed mixed-micelles with glycosphingolipids. The glycolipids were purified by DEAE-Sephadex A-25 column chromatography. The fractions containing mixed micelles were obtained from linear gradient solvent of 0.05M-0.5M ammonium acetate in methanol. HPLC ( Bondapak-NH(2) - linked to a Bondapak-C(18) column) chromatogram showed five peaks. Two possible structures for the fourth peak fraction were proposed as (CH(3))(3)C-CH(2)-C(CH(3))(2)-C(6)H(4)-OR and (CH(3))(3)C-C(CH(3))(2)-CH(2)-C(6)H(4)-OR by NMR spectroscopy. The structure was further confirmed by electrospray tandem mass spectrometry (ESI MS/MS). The spectrum showed a protonated molecule at m/z 559 and three different series of ions with mass difference of 44 were detected in the MS/MS spectrum. Therefore, the structure of the fourth peak fraction from HPLC was confirmed as octoxynol, (CH(3))(3)C-CH(2)-C(CH(3))(2)-C(6)H(4)-(OCH(2)-CH(2))n-OH, based on mass spectrometry and NMR spectroscopy.

Animals↗

Differential modulation by muscimol and baclofen on antinociception induced by morphine, beta-endorphin, D-Pen2,5-enkephalin and U50,488H administered intracerebroventricularly in the mouse.

The present study was designed to investigate the modulatory effects of stimulation of GABAA and GABAB receptors at supraspinal sites on antinociception induced by supraspinally administered mu-, epsilon-, delta-, and kappa-opioid receptor agonists. The effects of the GABAA and GABAB receptor agonists, muscimol and baclofen respectively, on the antinociception induced by morphine (a mu-receptor agonist), beta-endorphin (an epsilon-receptor agonist), D-Pen2,5-enkephalin (DPDPE, a delta-receptor agonist) and U50,488H ([trans-3,4-dichloroN-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl] benzeocetamide]; a kappa-receptor agonist) injected intracerebroventricularly (i.c.v.) were studied. The antinociception was assayed using the tail-flick and hot-plate tests. Muscimol at doses of 25-200 ng, administered i.c.v. alone did not affect the latencies of tail-flick and hot-plate thresholds, but attenuated dose-dependently the inhibition of the tail-flick and hot-plate responses induced by i.c.v. administered morphine (2 micrograms), beta-endorphin (1 microgram), DPDPE (10 micrograms), and U50,488H (60 micrograms). Baclofen (1.25-10 ng) administered i.c.v. alone did not affect the latencies of the tail-flick and hot-plate responses, but attenuated dose-dependently the inhibition of the tail-flick and hot-plate responses induced by beta-endorphin and U50,488H, without affecting morphine- or DPDPE-induced responses. Our results indicate that activation of GABAA receptors at the supraspinal sites by i.c.v. injection of muscimol antagonizes antinociception induced by supraspinally administered mu-, epsilon-, delta-, and kappa-opioid receptor agonists. On the other hand, activation of GABAB receptors at supraspinal sites by i.c.v. baclofen antagonizes antinociception induced by i.c.v. administered epsilon- and kappa-opioid agonists, but not mu- or delta-opioid agonists.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗

Non-NMDA receptor antagonist attenuates antinociception induced by morphine but not beta-endorphin, D-Pen2-D-Pen5-enkephalin, and U50, 488H administered intracerebroventricularly in mice.

The antinociception induced by morphine but not beta-endorphin, D-Pen2-D-Pen5-enkephalin (DPDPE), or U50, 488H (trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl) cyclohexyl] benzeocetamide) administered intracerebroventricularly (i.c.v.) has been previously demonstrated to be mediated by the N-Methyl-D-Aspartic Acid (NMDA) receptor. The present study was designed to determine if non-NMDA receptors are involved in opioid-induced antinociception. Antinociception was assessed by the tail-flick test in male ICR mice. Various doses of CNQX (6-Cyano-7-nitroquinoxaline-2,3-dione), a competitive non-NMDA receptor antagonist, (0.001 to 0.5 microgram) injected intracerebroventricularly (i.c.v.) alone did not show any antinociceptive effect. CNQX (0.01 to 1 micrograms, i.c.v.) dose-dependently attenuated the inhibition of the tail-flick response induced by morphine (1 microgram). However, inhibition of the tail-flick response induced by i.c.v. administered beta-endorphin (1 microgram), DPDPE (10 micrograms), or U50, 488H was not affected by i.c.v. administered CNQX. It is concluded that non-NMDA receptors are involved in i.c.v. morphine-induced antinociception. However, non-NMDA receptors are not involved in i.c.v. administered beta-endorphin-, DPDPE-, and U50, 488H-induced antinociception at the supraspinal level.

6-Cyano-7-nitroquinoxaline-2,3-dione↗

Differential antagonism by MK-801 against antinociception induced by opioid receptor agonists administered supraspinally in mice.

Various doses of MK-801 ((+/-)-5-methyl-10,11-dihydro-5H-dibenzo(a,d) cyclohepten-5, 10-imine maleate), a non-competitive N-methyl-D-aspartic acid (NMDA) receptor antagonist (0.001-1 microgram) injected intracerebroventricularly (i.c.v.) alone did not show any antinociceptive effect. MK-801 (0.001-1 microgram i.c.v.) dose dependently attenuated the inhibition of the tail-flick and hot plate responses induced by i.c.v. administered morphine (1 microgram), [D-Pen2, D-Pen5]enkephalin (DPDPE; 10 micrograms), and U50,488H (trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl]benzeoce tamide ) 60 micrograms). However, the inhibition of the tail-flick and hot plate responses induced by i.c.v. administered beta-endorphin (1 microgram) was not changed by i.c.v. administered MK-801. Our results indicate that, at the supraspinal level, NMDA receptors are involved in the production of antinociception induced by supraspinally administered morphine, DPDPE, and U50,488H but not beta-endorphin.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗

Role of three rab5-like GTPases, Ypt51p, Ypt52p, and Ypt53p, in the endocytic and vacuolar protein sorting pathways of yeast.

The small GTPase rab5 has been shown to represent a key regulator in the endocytic pathway of mammalian cells. Using a PCR approach to identify rab5 homologs in Saccharomyces cerevisiae, two genes encoding proteins with 54 and 52% identity to rab5, YPT51 and YPT53 have been identified. Sequencing of the yeast chromosome XI has revealed a third rab5-like gene, YPT52, whose protein product exhibits a similar identity to rab5 and the other two YPT gene products. In addition to the high degree of identity/homology shared between rab5 and Ypt51p, Ypt52p, and Ypt53p, evidence for functional homology between the mammalian and yeast proteins is provided by phenotypic characterization of single, double, and triple deletion mutants. Endocytic delivery to the vacuole of two markers, lucifer yellow CH (LY) and alpha-factor, was inhibited in delta ypt51 mutants and aggravated in the double ypt51ypt52 and triple ypt51ypt52ypt53 mutants, suggesting a requirement for these small GTPases in endocytic membrane traffic. In addition to these defects, the here described ypt mutants displayed a number of other phenotypes reminiscent of some vacuolar protein sorting (vps) mutants, including a differential delay in growth and vacuolar protein maturation, partial missorting of a soluble vacuolar hydrolase, and alterations in vacuole acidification and morphology. In fact, vps21 represents a mutant allele of YPT51 (Emr, S., personal communication). Altogether, these data suggest that Ypt51p, Ypt52p, and Ypt53p are required for transport in the endocytic pathway and for correct sorting of vacuolar hydrolases suggesting a possible intersection of the endocytic with the vacuolar sorting pathway.

Amino Acid Sequence↗