• 생명과학회지
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• 제13권1호
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• pp.67-72
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• 2003

위염, 위궤양 및 위암의 원인 균인 Helicobacter pylori 가균체 표면에 다량 함유하며 주된 생존 인자이며 병원성 인자인 urease를 대장균에서 발현시키고 이 효소에 대한 항체 또는 기질과의 특이 상호 작용을 이용하여 두 단계의 간편한 방법에 의하여 정제하였다. 우선 anti-H. pylori Urease IgG-Sepharose column과 urea-Sepharose column을 각각 제조하고 DEAE-Sepharose 음이온 교환수지를 통하여 1차 정제한 시료를 각각 적용하고 제반 조건에서 용출시켰다. Anti-H. pylori urease IgG-Sepharose column의 경우에는 urease 시료가 너무 강력하게 결합함으로써 극단적인 pH조건에서만 용출이 가능함이 관찰되었으므로, 100 mM 탄산 완충액(pH 10.5)으로 최종 용출하였을 때 비교적 순수한 효소를 얻었으나, 비활성이 다소 감소된 것으로 나타났다. 한편, urea-Sepharose에 적용시킨 시료는 100 mM urea-HEB 완충액(pH 7.5)으로 비교적 용이하게 용출되어 비교적 높은 순도와 비활성의 urease 효소를 얻을 수 있었으나 이 경우에는 urease의 smaller subunit인 UreA peptide band의 강도가 다소 감소한 것이 관찰되었다.

• 생약학회지
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• 제45권2호
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• pp.168-173
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• 2014

The aim of this study was to evaluate the anti-helicobacter activity of the ethanol extract of Sohamhyoongtang (Coptidis Rhizoma, Pinelliae Tuber and Trichosanthis Semen) and Coptidis Rhizoma total alkaloids, which is one of the components of Sohamhyoongtang. Crude ethanol extract of Sohamhyoongtang (ESHHT) and Coptidis Rhizoma total alkaloids (CRTA) were used for this experiment. Five different types of H. pylori (including H. pylori 26695) were used as test strain. To determine anti-helicobacter activity, minimum inhibitory concentration (MIC) was determined by agar dilution method. The effect of ESHHT and CRTA on the gene expression of H. pylori was investigated by quantitative realtime-PCR (qRT-PCR). MICs of ESHHT against five H. pylori strains were $250{\sim}500{\mu}g/ml$ and MICs of CRTA against five H. pylori strains were $50{\sim}200{\mu}g/ml$. Four representative virulence genes of H. pylori, cagA, ureA, ureB and ureI were tested as target genes for qRT-PCR. According to the qRT-PCR results, both ESHHT and CRTA markedly repressed the expression of cagA gene of H. pylori 26695 (6.91 and 20 folds respecively). These results showed that the ESHHT and CRTA demonstrated antihelicobacter properties, suggesting their potential use in gastritis or duodenal ulcer.

• 한국임상수의학회지
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• 제24권3호
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• pp.305-307
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• 2007

락토페린 결합단백질(Lactoferrin-binding proteins, LBP)은 젖소유방염 원인균인 Streptococcus uberis의 막단백질로서 그 특성에 관해서는 잘 규명되어 있지 않지만, 특히 최근에는 스트렙토코커스성 유방염의 독성인자로서 중요시되고 있다. 본 연구에서는 S. uberis 네 가지 균주를 대상으로 LBP를 보다 효율적으로 추출하기 위하여 mutanolysin 및 sodium dodecyl sulfate(SDS)를 이용한 두 가지 다른 추출 방법을 사용하였다. 추출된 세균단백질을 SDS-polyacrylamide gel electrophoreis(SDS-PAGE)로 전기영동을 하였고, 겔을 니트로셀룰로스 막으로 이동시켰다. Rabbit anti-bovine lactoferrin 항체와 HRP-conjugated donkey anti-rabbit IgG 항체를 사용하여 LBP를 검출하였다. 이러한 웨스턴 블롯팅 분석을 통해 SDS 추출법이 mutanolysin 추출법에 비해 보다 효율적으로 110 kDa 및 112 kDa의 LBP를 추출할 수 있음을 증명하였다.

• Genomics & Informatics
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• 제20권4호
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• pp.47.1-47.13
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• 2022

Klebsiella pneumoniae is a gram-negative bacterium that is known for causing infection in nosocomial settings. As reported by the World Health Organization, carbapenem-resistant Enterobacteriaceae, a category that includes K. pneumoniae, are classified as an urgent threat, and the greatest concern is that these bacterial pathogens may acquire genetic traits that make them resistant towards antibiotics. The last class of antibiotics, carbapenems, are not able to combat these bacterial pathogens, allowing them to clonally expand antibiotic-resistant strains. Most antibiotics target essential pathways of bacterial cells; however, these targets are no longer susceptible to antibiotics. Hence, in our study, we focused on a hypothetical protein in K. pneumoniae that contains a DNA methylation protein domain, suggesting a new potential site as a drug target. DNA methylation regulates the attenuation of bacterial virulence. We integrated computational-aided drug design by using a bioinformatics approach to perform subtractive genomics, virtual screening, and fingerprint similarity search. We identified a new potential drug, koenimbine, which could be a novel antibiotic.

• IMMUNE NETWORK
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• 제20권5호
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• pp.37.1-37.15
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• 2020

Mycobacterium tuberculosis (Mtb) is an etiologic pathogen of human tuberculosis (TB), a serious infectious disease with high morbidity and mortality. In addition, the threat of drug resistance in anti-TB therapy is of global concern. Despite this, it remains urgent to research for understanding the molecular nature of dynamic interactions between host and pathogens during TB infection. While Mtb evasion from phagolysosomal acidification is a well-known virulence mechanism, the molecular events to promote intracellular parasitism remains elusive. To combat intracellular Mtb infection, several defensive processes, including autophagy and apoptosis, are activated. In addition, Mtb-ingested phagocytes trigger inflammation, and undergo necrotic cell death, potentially harmful responses in case of uncontrolled pathological condition. In this review, we focus on Mtb evasion from phagosomal acidification, and Mtb interaction with host autophagy, apoptosis, and necrosis. Elucidation of the molecular dialogue will shed light on Mtb pathogenesis, host defense, and development of new paradigms of therapeutics.

• 대한약침학회지
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• 제26권4호
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• pp.307-318
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• 2023

Objectives: Bacterial biofilm is regarded as a significant threat to the production of safe food and the arise of antibiotic-resistant bacteria. The objective of this investigation is to evaluate the quorum sensing inhibitory effect of Nepeta curviflora methanolic extract. Methods: The effectiveness of the leaves at sub-inhibitory concentrations of 2.5, 1.25, and 0.6 mg/mL on the virulence factors and biofilm formation of P. aeruginosa was evaluated. The effect of N. curviflora methanolic extract on the virulence factors of P. aeruginosa, including pyocyanin, rhamnolipid, protease, and chitinase, was evaluated. Other tests including the crystal violet assay, scanning electron microscopy (SEM), swarming motility, aggregation ability, hydrophobicity and exopolysaccharide production were conducted to assess the effect of the extract on the formation of biofilm. Insight into the mode of antiquorum sensing action was evaluated by examining the effect of the extract on the activity of N-Acyl homoserine lactone (AHL) and the expression of pslA and pelA genes. Results: The results showed a significant attenuation in the production of pyocyanin and rhamnolipid and in the activities of protease and chitinase enzymes at 2.5 and 1.25 mg/mL. In addition, N. curviflora methanolic extract significantly inhibited the formation of P. aeruginosa biofilm by decreasing aggregation, hydrophobicity, and swarming motility as well as the production of exopolysaccharide (EPS). A significant reduction in AHL secretion and pslA gene expression was observed, indicating that the extract inhibited quorum sensing by disrupting the quorum-sensing systems. The quorum-sensing inhibitory effect of N. curviflora extract appears to be attributed to the presence of kaempferol, quercetin, salicylic acid, rutin, and rosmarinic acid, as indicated by LCMS analysis. Conclusion: The results of the present study provide insight into the potential of developing anti-quorum sensing agents using the extract and the identified compounds to treat infections resulting from quorum sensing-mediated bacterial pathogenesis.

• 한국식품위생안전성학회지
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• 제39권3호
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• pp.231-238
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• 2024

국내에서 V. parahaemolyticus로 인한 식중독 사고가 지속적으로 보고되고 있으며, 최근 국내 수산물 판매량 및 수산물 양식에 사용되는 항생제 판매량은 증가하는 추세이다. 따라서 본 연구는 국내에 유통되는 수산물에서 분리한 V. parahaemolyticus의 분포, 항생제 감수성, 유전적 특성 및 유전학적 통계를 조사하였다. 79건의 유통 수산물로부터 47건(59.5%)에서 V. parahaemolyticus가 분리되었다. 항생제 내성 양상의 경우, 총 47균주의 분리 균주에서는 ampicillin에 2균주(4.3%)가 내성을 보였으며, 이외 균주는 모든 항생제에 대해 감수성을 보였다. 항생제 내성 유전자의 경우, 모든 균주(100%)로부터 bla_CAR family gene, tet(35), catC가 확인되었으며, 1균주(2.1%)에서는 fos가 확인되었다. 병원성 유전자 여부의 경우, 모든 분리 균주에서 tdh, trh 유전자는 확인되지 않았으나, T3SS1은 모든 균주(100%), T3SS2는 1균주(2.1%)에서 확인되었다. MLST의 경우, 17균주로부터 15가지의 ST가 확인되었으며, ST 658가 3균주, 이외 14가지 ST는 1균주씩 확인되었다. 확인된 ST는 대부분 중국, 태국 등의 환경 분리주로 확인되었으며, ST 396, ST 3042는 중국 임상 분리주로부터 확인되었다. 이로써, 최근 국내에 수산물과 관련한 식중독, 유통량, 항생제 판매량 등의 추세에 따른 위험성에 V. parahaemolyticus에 대한 지속적인 연구가 필요할 것으로 사료되며, 본 연구는 그에 대한 도움이 될 것이라 사료된다.

• Journal of Microbiology and Biotechnology
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• 제27권10호
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• pp.1844-1854
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• 2017

Trichomonas vaginalis is a pathogen that triggers severe immune responses in hosts. T. vaginalis ${\alpha}$-actinin 2 ($Tv{\alpha}$-actinin 2) has been used to diagnose trichomoniasis. $Tv{\alpha}$-actinin 2 was dissected into three parts; the N-terminal, central, and C-terminal portions of the protein (#1, #2, and #3, respectively). Western blot of these $Tv{\alpha}$-actinin 2 proteins with pooled patients' sera indicated that #2 and #3, but not #1, reacted with those sera. Immunofluorescence assays of two different forms of T. vaginalis (trophozoites and amoeboid forms), using anti-$Tv{\alpha}$- actinin 2 antibodies, showed localization of $Tv{\alpha}$-actinin 2 close to the plasma membranes of the amoeboid form. Fractionation experiments indicated the presence of $Tv{\alpha}$-actinin 2 in cytoplasmic, membrane, and secreted proteins of T. vaginalis. Binding of fluorescence-labeled Trichomonas to vaginal epithelial cells and prostate cells was decreased in the antibody blocking experiment using anti-$Tv{\alpha}$-actinin 2 antibodies. Pretreatment of T. vaginalis with anti-$rTv{\alpha}$-actinin 2 antibodies also resulted in reduction in its cytotoxicity. Flow cytometry, ligand-binding immunoblotting assay, and observation by fluorescence microscopy were used to detect the binding of recombinant $Tv{\alpha}$-actinin 2 to human epithelial cell lines. Specifically, the truncated N-terminal portion of $Tv{\alpha}$-actinin 2, $Tv{\alpha}$-actinin 2 #1, was shown to bind directly to vaginal epithelial cells. These data suggest that ${\alpha}$-actinin 2 is one of the virulence factors responsible for the pathogenesis of T. vaginalis by serving as an adhesin to the host cells.

• 대한본초학회지
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• 제24권4호
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• pp.39-47
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• 2009

Objectives : Hwangnyeonhaedok-tang (Huanglian Jiedu Tang; HHT) has been widely used for purging' 'fire' and lessening virulence of any pathogenic organism. However it has been rarely conducted to evaluate the immuno-biological activity. In this study, we evaluated anti-inflammatory effects of HHT in LPS-activated Raw264.7 cells. Methods : Cells were treated with $1\;{\mu}g/ml$ of LPS 1 h prior to the addition of HHT. Cell viability was measured by MTT assay. The production of NO was determined by reacting cultured medium with Griess reagent. PGE2 and proinflammatory cytokines were detected by ELISA. Expression of iNOS, COX-2, $I{\kappa}B{\alpha}$ and NF-${\kappa}B$ were analyzed by immunoblot analysis. Results : All three doses of HHT (0.03, 0.10 and 0.30 mg/ml) had no significant cytotoxicity during the entire experimental period. The levels of NO and PGE2 were dramatically augmented by LPS compared to control. However, HHT extract dose-dependently reduced these increases. Expression of iNOS and COX-2 protein were also decreased by treatment with HHT extract. Furthermore, HHT extract significantly reduced the nuclear translocation of NF-${\kappa}B$ which is critical in regulating inflammation through transcription of iNOS and COX-2. In addition, HHT extract reduced the elevated production of inflammatory cytokines including TNF-$\alpha$, IL-$1{\beta}$ and IL-6. Conclusions : The results in this study demonstrate that HHT extract exerts anti-inflammatory activities through the inhibition of NO, PGE2 and proinflammatory cytokines production via the suppression of NF-${\kappa}B$.

• 대한화장품학회지
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• 제50권1호
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• pp.19-27
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• 2024

본 연구는 배롱나무 꽃(Lagerstroemia indica flower)을 항여드름 화장품 소재로 제안하기 위한 것으로 배롱나무 꽃의 에탄올 추출물(Lagerstroemia indica flower extract, LIFE)을 이용해 피지조절과 항염, 항산화 효능을 확인하였다. 피지조절 효능을 평가한 결과 LIFE가 팔미트산으로 과도하게 유도된 피지생성을 65%까지 억제함을 확인하였다. 이어서 LIFE가 여드름 병변에서 주로 나타나는 Cutibacterium acnes (C. acnes)에 대한 항균작용과 lipase 활성억제능이 있으며, C. acnes로부터 분비되는 독성인자에 의한 염증반응도 감소시킬 수 있음을 확인하였다. 또한 LIFE가 대식세포에서 LPS로 유도된 염증성 매개인자인 일산화질소(NO)와 prostaglandin E₂ (PGE₂)의 분비를 각각 75%, 54%까지 억제함을 확인하였으며, 아스코르브산과 비슷한 정도의 높은 DPPH 라디칼 소거능 또한 있음을 확인하였다. 이 결과들은 천연 추출물인 LIFE가 피지 생성 조절과 함께, C. acnes에 대한 항균 및 lipase 활성 억제, 항염, 항산화능을 가짐으로써 복합성 질환인 여드름을 완화 및 치료하는 항여드름 소재로 활용될 수 있음을 제시한다.