• BMB Reports
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• 제44권6호
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• pp.405-409
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• 2011

The present study demonstrated that valproic acid (VPA) transcriptionally regulates human GM3 synthase (hST3Gal V), which catalyzes ganglioside GM3 biosynthesis in ARPE-19 human retinal pigment epithelial cells. For this, we characterized the promoter region of the hST3Gal V gene. Functional analysis of the 5'-flanking region of the hST3Gal V gene revealed that the -177 to -83 region functions as the VPA-inducible promoter and that the CREB/ATF binding site at -143 is crucial for VPA-induced expression of hST3Gal V in ARPE-19 cells. In addition, the transcriptional activity of hST3Gal V induced by VPA in ARPE-19 cells was inhibited by SP600125, a c-Jun N-terminal kinase (JNK) inhibitor. In summary, our results identified the core promoter region in the hST3Gal V promoter and for the first time demonstrated that ATF2 binding to the CREB/ATF binding site at -143 is essential for transcriptional activation of hST3Gal V in VPA-induced ARPE-19 cells.

• Journal of Periodontal and Implant Science
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• 제39권sup2호
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• pp.231-237
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• 2009

Purpose: Cytolethal distending toxin (CDT) is a family of heat-labile cytotoxins produced by several gram-negative mucosa-associated pathogens, including Aggregatibacter actinomycetemcomitans. CDT is well known to be capable of inducing growth arrest, morphological alterations, and eventually death in various cells. CDT belongs to a tripartite $AB_2$ toxin (CdtB: the enzymatic A subunit; CdtA and CdtC: the heterodimeric B subunit). Previous studies proposed that CdtA and CdtC together bind to a cell surface receptor and glycolipids act as a receptor for A. actinomycetemcomitans CDT (AaCDT). In this study, recombinant CdtA and CdtC proteins of AaCDT were co-expressed in a bacterial expression system and tested for their affinity for $GM_1$ ganglioside. Methods: The genes for CdtA and CdtC from A. actinomycetemcomitans Y4 were utilized to construct the expression vectors, pRSET-cdtA and pET28a-cdtC. Both CdtA and CdtC proteins were expressed in Escherichia coli BL21(DE3) and then purified using hexahistidine (His6) tag. The identity of purified protein was confirmed by anti-His6 antibody and monoclonal anti-CdtA antibody. Furthermore, the affinity of recombinant protein to $GM_1$ ganglioside was checked through ELISA. Results: Recombinant CdtA and CdtC proteins were expressed as soluble proteins and reacted to anti-His6 and monoclonal anti-CdtA antibodies. ELISA revealed that purified soluble CdtA-CdtC protein bound to $GM_1$ ganglioside, while CdtA alone did not. Conclusions: Co-expression of CdtA and CdtC proteins enhanced the solubility of the proteins in E. coli, leading to convenient preparation of active CdtA-CdtC, a critical material for the study of AaCDT pathogenesis.

• 한국당과학회:학술대회논문집
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• 한국당과학회 2006년도 제1회 연례학술대회
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• pp.28-29
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• 2006

• Molecules and Cells
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• 제20권3호
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• pp.354-360
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• 2005

Neuronal damage subsequent to transient cerebral ischemia is a multifactorial process involving several overlapping mechanisms. Gangliosides, sialic acid-conjugated glycosphingolipids, reduce the severity of acute brain damage in vitro. However their in vivo effects on the cerebral cortex damaged by ischemic infarct are unknown. To assess the possible protective role of gangliosides we examined their expression in the cerebral cortex damaged by ischemic infarct in the rat. Ischemia was induced by middle cerebral artery (MCA) occlusion, and the resulting damage was observed by staining with 2, 3, 5-triphenylterazolium chloride (TTC). High-performance thin-layer chromatography (HPTLC) showed that gangliosides GM3 and GM1 increased in the damaged cerebral cortex, and immunofluorescence microscopy also revealed a significant change in expression of GM1. In addition, in situ hybridization demonstrated an increase in the mRNA for ganglioside GM3 synthase. These results suggest that gangliosides GM1 and GM3 may be synthesized in vivo to protect the cerebral cortex from ischemic damage.

• Molecules and Cells
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• 제37권2호
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• pp.161-171
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• 2014

In several lysosomal storage disorders, including Niemann-Pick disease Type C (NP-C), sphingolipids, including glycosphingolipids, particularly gangliosides, are the predominant storage materials in the brain, raising the possibility that accumulation of these lipids may be involved in the NP-C neurodegenerative process. However, correlation of these accumulations and NP-C neuropathology has not been fully characterized. Here we derived NP-C mice with complete and partial deletion of the Siat9 (encoding GM3 synthase) gene in order to investigate the role of ganglioside in NP-C pathogenesis. According to our results, NP-C mice with homozygotic deletion of GM3 synthase exhibited an enhanced neuropathological phenotype and died significantly earlier than NP-C mice. Notably, in contrast to complete depletion, NP-C mice with partial deletion of the GM3 synthase gene showed ameliorated NP-C neuropathology, including motor disability, demyelination, and abnormal accumulation of cholesterol and sphingolipids. These findings indicate the crucial role of GM3 synthesis in the NP-C phenotype and progression of CNS pathologic abnormality, suggesting that well-controlled inhibition of GM3 synthesis could be used as a therapeutic strategy.

• 한국수정란이식학회지
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• 제32권1호
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• pp.17-24
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• 2017

Ganglioside GD1a is specifically formed by the addition of sialic acid to ganglioside GM1a by ST3 ${\beta}$-galactoside ${\alpha}$-2,3-sialyltransferase 2 (ST3GAL2). Above all, GD1a are known to be related with the functional regulation of several growth factor receptors, including activation and dimerization of epidermal growth factor receptor (EGFR) in tumor cells. The activity of EGF and EGFR is known to be a very important factor for meiotic and cytoplasmic maturation during in vitro maturation (IVM) of mammalian oocytes. However, the role of gangliosides GD1a for EGFR-related signaling pathways in porcine oocyte is not yet clearly understood. Here, we investigated that the effect of ST3GAL2 as synthesizing enzyme GD1a for EGFR activation and phosphorylation during meiotic maturation. To investigate the expression of ST3GAL2 according to the EGF treatment (0, 10 and 50 ng/ml), we observed the patterns of ST3GAL2 genes expression by immunofluorescence staining in denuded oocyte (DO) and cumulus cell-oocyte-complex (COC) during IVM process (22 and 44 h), respectively. Expression levels of ST3GAL2 significantly decreased (p<0.01) in an EGF concentration (10 and 50 ng/ml) dependent manner. And fluorescence expression of ST3GAL2 increased (p<0.01) in the matured COCs for 44 h. Under high EGF concentration (50 ng/ml), ST3GAL2 protein levels was decreased (p<0.01), and their shown opposite expression pattern of phosphorylation-EGFR in COCs of 44 h. Phosphorylation of EGFR significantly increased (p<0.01) in matured COCs treated with GD1a for 44 h. In addition, ST3GAL2 protein levels significantly decreased (p<0.01) in GD1a ($10{\mu}M$) treated COCs without reference to EGF pre-treatment. These results suggest that treatment of exogenous ganglioside GD1a may play an important role such as EGF in EGFR-related activation and phosphorylation in porcine oocyte maturation of in vitro.

• Bulletin of the Korean Chemical Society
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• 제19권5호
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• pp.569-575
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• 1998

Investigation of the structure of the gangliosides has proven to be very important in the understanding of their biological roles. We have determined the tertiary structure of asialoganglioside GM1 $(GA_1)$ using NMR spectroscopy and distance geometry calculations. All of the structures are very similar except the glycosidic torsion angles in the ring IV and ring III linkages. There are two low-energy structures for GA1, G1 and G2. G1 differs from G2 only in the IV-III glycosidic linkages and the orientation of acetamido group in ring III. There is a stable intramolecular hydrogen bond between the third hydroxyl group in ring I and the ring oxygen atom in ring II. Also, there may be a weak hydrogen bond between the second hydroxyl group in ring IV and the acetamido group in ring III. Small coupling constants of $^3J_{IH3,IOH3}\; and\; ^3J_{IVH2,IVOH2}$ support this result. Overall structural features of $(GA_1)$ are very similar to those of $(GM_1)$. It implicates that specificities of the sugar moieties in GM1 are caused not by their tertiary foldings, but mainly by the electrostatic interactions between the polar sialic acid and its receptors. Since it is evident that $(GA_1)$ is more hydrophobic than $(GA_1)$, a receptor with a hydrophobic binding site can recognize the $(GA_1)$ better than $(GA_1)$. Studies on the conformational properties of $(GA_1)$ may lead to a better understanding of the molecular basis of its functions.

• 한국진공학회:학술대회논문집
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• 한국진공학회 2016년도 제50회 동계 정기학술대회 초록집
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• pp.378.1-378.1
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• 2016

나노바이오연구분야에서 ToF-SIMS를 이용하여 lipid와 metabolite같은 저 분자의 생체물질을 측정하는데 널리 이용되어 왔다. 최근에는 고 분자량의 생체물질을 측정하기 위해서 C60, water cluster, argon cluster등의 다양한 종류의 클러스터 이온빔들이 개발되어 왔다. [1,2] 하지만 tissue샘플을 클러스터 이온빔을 이용하여 분석한 결과에서도 m/z 1500이상의 고분자를 측정한 결과는 거의 없다. 바이오샘플의 charging을 상쇄하기위해 low energy electron beam (~20 eV)을 사용하는데, low energy electron beam이 샘플에 damage를 주기 때문이다. [3] 본 연구에서는 electron fluence (electrons/cm2)가 증가함에 따라 PC(16:0/18:1(9Z)와 Ganglioside GM1의 intensity가 감소함을 알았고, low energy electron beam에 의해 생체 물질이 damage를 받을 수 있음을 확인하였다. 따라서 tissue 샘플을 SUS기판에 샘플링하고 Ar-GCIB를 이용하면 charging없이 tissue imaging을 성공적으로 수행할 수 있고, m/z 2000이상의 고 분자량의 생체물질을 측정할 수 있음을 확인하였다.

• 한국축산식품학회지
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• 제40권2호
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• pp.209-220
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• 2020

Milk fat globule membrane (MFGM) is a lipid carrier in mammals including humans that consists mainly of polar lipids, like phospholipids and glycolipids. In this study, a process to enrich polar lipids in commercial butter and whey powder, including polar lipids of MFGM, was developed. WPC (whey protein concentrate) 60 was selected as the most suitable raw material based on the yield, phospholipid, protein, and lactose content of the polar lipid fraction obtained by ethanol extraction of two WPC (WPC60 and WPC70) and two buttermilk (A and B). After fractionation under optimum conditions, the polar-lipid enriched fraction from WPC60 contained 38.56% phospholipids. The content of glycolipids, cerebroside, lactosylceramide, ganglioside GM3, ganglioside GD3, was 0.97%, 0.55%, 0.09%, and 0.14%, respectively. Rancimat results showed that the oxidation stability of fish oil increased with an increase in the polar-lipid fraction by more than 30 times. In addition, the secretion of IL-6 and TNF-α decreased in a concentration-dependent manner after treatment of RAW 264.7 cells with 0.1 to 100 ppm of the polar lipid fraction. In this study, polar lipid concentrates with antioxidant and anti-inflammatory activity, were prepared from milk processing by-products. The MFGM polar lipid concentrates made from by-products are not only additives for infants, but are also likely to be used as antioxidants in cooking oils and as active ingredients for functional foods.

• 한국동물위생학회지
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• 제20권4호
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• pp.359-370
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• 1997

This study was carried out to Investigate the biochemical characteristics, antibiotic susceptibility, serogroups and pili producibility test of enterotoxigenic Escherichia coli(ETEC) isolated from piglets with diarrhea in Kangwon province from March to October 1996. 1. Sixty eight E coli strains were isolated from 72 piglets with diarrhea and the biochemical and cultural reaction were compared with the classification criteria of Edwards and Ewing. 2. The serogroups of 26 isolates were classified as 08 : K87 6(8.8%), O20 : K1O1 4(5.9%), O141 : K85 4(5.9%), 09 : K103 : P987 3(4.4%), O45 : K 2(2.9%) 0139 : K82 2(2.9%), O64 : $K^{-}$2(2.9%), O149 : K91 1(1.5%), O157 : K88ac 1(1.5%) and O115 : $K^{-}$1(1.5%), respectively. 3. In antibiotic susceptibility test, the isolates showed high susceptible to Ak, Eno, Na, Gm, Am and Km, whereas resistance to Tc, Sm and Cf. 4. Sixty one strains(89.7%) of 68 I coli Isolates were resistant to one or more drugs. The isolates resistant to 2 and 3 or more drugs were 60.3% and 19.1%, respectively. Amog the 16 multiple resistant patterns, Sm Tc(11.5% ), Cf Sm Tc(11.5% ), Cf Cp Sm Su Tc(9.8% ) and Cf Cp Sm Su Tc(8.2%) patterns were frequently observed. 5. MRHA of guinea pig erythrocytes was detected in 9 out of 26 OK serotype and 9 out of 42 unidentified serotypes. MRHA titers of serotypes showed from 16 to 32 in O141 : K85 and no titers in O139 : K82. 6. By the GM1 ganglioside ELISA, $\beta$-, $\alpha$-, and $\gamma$-hemolysin producing strains was detected as 36, 6, and 5 from heat labile enterotoxin(LT) of 47 ETEC, respectively. The distribution of LT toxin from 112 isolates was showed $\beta$- hemolysin, 2 isolates $\alpha$-hemolysin and 3 isolates $\gamma$-hemol-ysin from 26 OK serotypes.