• The Korean Journal of Physiology and Pharmacology
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• 제8권3호
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• pp.141-146
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• 2004

Soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNARE) proteins, composed of two presynaptic membrane proteins [synaptosomal-associated protein of 25 kDa (SNAP-25) and syntaxin] and a presynaptic vesicular protein [vesicle-associated membrane protein (VAMP)], serve as a core of exocytotic fusion machinery, which can be affected by ischemia. Synaptic protein in core region, striatum and cortex has been shown to alter after focal ischemia, however, little is known in hippocampus. Hippocampus is remote from ischemic core, but it is one of the most vulnerable regions. Using immunohistochemistry, the present study was undertaken to investigate the alteration of expression of SNAP-25, syntaxin, and VAMP in the hippocampus of rats which were subjected to middle cerebral artery occlusion (MCAO) for 2h and allowed to reperfuse. At 2 weeks of reperfusion, the SNAP-25 and syntaxin immunoreactivity was increased in the stratum oriens of the CA1 and the stratum lucidum of the CA3 in the ipsilateral hippocampus. However, VAMP immunoreactivity didn't show significant change. These results demonstrate that the level of the presynatpic plasma membrane proteins (SNAP-25 and syntaxin) in the rat hippocampus is more sensitively affected by focal ischemia than that of the synaptic vesicle protein (VAMP).

• KSBB Journal
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• 제19권6호
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• pp.462-466
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• 2004

S. coelicolor A3(2)로부터 항산화 저분자 thiol분자인 MSH를 HPLC 및 monobromobimane 형광 검출 방법으로 분리${\cdot}$정제하여 그 존재를 확인하였다. 표준물질인 MSH-bimane과 동일하게 용출되는 MSH 분획을 확인하였으며 여러 thiol 분획 중 MSH 분획이 가장 많은 것으로 보아 MSH가 S. coelicolor의 주된 thiol 화합물로 판단되었다. MSH 생합성에 관여하는 효소 중 I-1-Pase의 유전자의 기능을 알아보기 위하여 이 유전자를 방선균에서 분리한 후 대장균에 클로닝하여 과도발현시켰다. 발현된 I-1-Pase를 Ni-NTA column을 사용하여 정제하였다. 정제된 I-1-Pase는 soluble protein으로 281개 아미노산으로 구성되어 있으며 분자량은 32 kDa이었다. 인간 및 대장균의 I-1-Pase와 각각 24와 $25\%$의 sequence homology를 보였으며, 기존의 I-1-Pase가 가지고 있는 공통의 I-1-Pase motif A와 motif B를 S. coelicolor A3(2)도 가지고 있는 것으로 확인되었다.

• 한국미생물·생명공학회지
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• 제42권2호
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• pp.190-193
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• 2014

CMP-Neu5Ac synthetase는 sialyated 된 glycoconjugates의 전구체로 사용되는 CMP-Neu5Ac를 합성하는데 관여하는 주요 효소이다. Escherichia coli K1에서 유래한 CMP-Neu5Ac synthetase 유전자 (neuA)는 평소 E. coli BL21(DE3)에서 비수용성으로 생성되는데, 이를 수용성 단백질로 생산하고자 여러 가지 샤페론 단백질 동시 발현기술을 이용하였다. 이를 위해, GroEL-ES와 DnaK-DnaJ-GrpE를 암호화하는 pG-KJE8 plasmid와 neuA를 동시 형질전환 시켰고 0.01 mM IPTG와 0.005 mg/ml의 L-arabinose로 유도하여 $20^{\circ}C$에서 발현시켰다. 그 결과, E. coli에서의 수용성 CMP-Neu5Ac Synthetase 생산이 현저하게 증가하였다.

• Journal of Plant Biotechnology
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• 제38권2호
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• pp.176-185
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• 2011

벼는 세계에서 가장 중요한 작물이며 크기가 383Mb로 게놈연구 모델 작물로 이용되고 있다. 또한 그 종자는 인간에게 탄수화물과 단백질 영양원을 제공한다. 벼 종자의 단백질은 약 8%를 차지하며 40%를 차지하는 콩 종자의 단백질 양에 비하여 상대적으로 적은 양을 나타낸다. 오스본의 분류에 의하면 종자 단백질은 수용성의 albumin, 염용해성 globulin, 알코올 용해성 prolamin 그리고 약산 또는 알카리 용해성 glutelin으로 나누어진다. Glutelin과 prolamin은 벼의 주요 저장단백질이다. 벼 glutelin 저장단백질 유전자의 발현분석을 위하여 일품벼 미숙종자의 발현유전자 (EST) 분석을 행하였다. 그 결과 11종의 미숙종자 발현 glutelin 유전자를 분리 하였으며 8개의 유전자는 염색체 2번에 위치하였다. Glutelin 유전자 발현양은 전체 미숙종자 발현유전자의 약 28.2%를 차지하였다. 또한 glu-04의 경우 같은 염색체 상에서 4.5 kb 떨어진 곳에 역방향의 같은 염기서열로 복제되어 있었다. 이와 같은 결과는 glutelin 유전자는 진화학적으로 복제되어 염색체 특이적으로 발현하는 것을 나타낸다. 종자 11개 glutelin 유전자들의 아미노산서열분석을 통하여 lysin 함량을 조사한 결과 glu05-type B7에서 4.51%의 높은 lysin 함량을 나타내었다. 향후 유전자의 과발현체를 이용한 lysin 함량을 높이는 영양성 강화 연구가 요구되어진다.

• 생명과학회지
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• 제17권1호
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• pp.132-136
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• 2007

E. coli에서 Pseudoalteromonas elyakovii 유래의 alginate lyase유전자(aly)를 발현시킬 때, 대부분의 단백질이 불용성 내포체 형태로 발현됨을 확인하였다. Alginate lyase를 가용성 활성형으로 생산하기 위해 aly와 DnaK/DnaJ/GrpE 또는 aly와 GroEL/ES을 공발현하는 형질전환체를 얻었다. 공발현 결과, 단백질의 올바른 접힘을 도와주는 DnaK/DnaJ/GrpE chaperone이 가용성 및 활성형의 alginate lyase 생산에 매우 효과적임을 알 수 있었다. DnaK/DnaJ/GrpE chaperone의 발현에 유도제인 L-arabinose 최적 농도는 0.05 mg/ml이었으며, 이러한 공발현에 의해 약 34%의 alginate lyase가 가용성 분획에서 생산되었다. 또한 10%의 cetylpyridinium chloride를 첨가함으로써, 공발현 콜로니 주위에 투명환이 형성됨을 확인할 수 있었고, 이는 활성형 alginate lyase 효소에 의해 alginate가 분해되었음을 시사하였다.

• Archives of Pharmacal Research
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• 제26권12호
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• pp.1061-1066
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• 2003

The fruit of Actinidia polygama (AP) has long been used as a folk medicine in Korea for treating pain, rheumatic arthritis and inflammation. The present investigation was carried out to determine the in vivo and in vitro anti-inflammatory activity of AP using several animal models of inflammation. The 70% ethanol extract of the fruit of AP significantly inhibited acetic acidinduced, vascular permeability in a dose dependent manner (23%, 38%, and 41 % inhibition at doses of 200 mg/kg, 500 mg/kg and 1000 mg/kg, respectively). This effect was maintained in AP water-soluble fraction (APW). The APW fraction also showed significant inhibitory activity against the rat paw edema induced by a single treatment of carrageenan. In vitro experiments were performed to demonstrate the inhibitory activities of APW (100 $\mu$ g/ml) on lipopolysaccharide (LPS)-induced nitric oxide (NO) and prostaglandin $E_2$ ($PGE_2$) production. The results showed that APW dose-dependently suppressed LPS-induced NO production in RAW 264.7 macrophages without a notable cytotoxic effect and also decreased inducible NO synthase (iNOS) protein expression. APW also showed a significant inhibitory effect in LPS-induced $PGE_2$ production and cyclooxygenase-2 (COX-2) expression.

• Molecules and Cells
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• 제21권1호
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• pp.7-20
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• 2006

The major event in human immunodeficiency virus type 1 (HIV-1) infection is the death of many cells related to host immune response. The demise of these cells is normally explained by cell suicide mechanism, apoptosis. Interestingly, the decrease in the number of immune cells, such as non-CD4+ cells as well as CD4+ T cells, in HIV infection usually occurs in uninfected bystander cells, not in directly infected cells. It has, therefore, been suggested that several soluble factors, including viral protein R (Vpr), are released from the infected cells and induce the death of bystander cells. Some studies show that Vpr interacts directly with adenine nucleotide translocator (ANT) to induce mitochondrial membrane permeabilization (MMP). The MMP results in release of some apoptogenic factors such as cytochrome-c (cyt-c) and apoptosis-inducing factor (AIF). Vpr also has indirect effect on mitochondria through enhancing the level of caspase-9 transcription and suppressing nuclear factor-kappa B (NF-${\kappa}B$). The involvement of p53 in Vpr-induced apoptosis remains to be studied. On the other hand, low level of Vpr expression has anti-apoptotic effect, whereas it's high level of expression induces apoptosis. Extracellular Vpr also exhibits cytotoxicity to uninfected bystander cells through apoptotic or necrotic mechanism. The facts that Vpr has cytotoxic effect on both infected cells and bystander cells, and that it exhibits both proand anti-apoptotic activity may explain its role in viral survival and disease progression.

• 대한의생명과학회지
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• 제30권3호
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• pp.113-122
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• 2024

In this study, we aimed to investigate the anti-cancer effects of the Pandanus tectorius extract on AGS cells. P. tectorius, commonly known as hala or screw pine, is a tropical plant traditionally used for its medicinal properties, including anti-inflammatory and antioxidant properties. Here, effects of the P. tectorius extract on cell proliferation, migration, and gene expression were evaluated using various assays, including the water-soluble tetrazolium salt (WST)-1, wound healing, migration, and western blotting assays. WST-1 assay revealed a significant dose- and time-dependent decrease in cell viability, with higher concentrations of the extract resulting in more pronounced viability inhibition. Wound healing and migration assays revealed that the P. tectorius extract effectively hindered cell migration, as the treated cells showed considerably slower wound closure and reduced migration than the control cells. Molecular analysis revealed that the extract significantly downregulated the expression levels of key oncogenic proteins, genes, and components of the Notch signaling pathway. Western blotting confirmed the substantial reduction in the marker protein levels in treated cells. These findings suggest that P. tectorius extract exerts its anti-cancer effects by inhibiting multiple signaling pathways crucial for cancer cell proliferation, migration, and survival. Overall, this study highlights the potential of P. tectorius extract as a therapeutic agent for gastric cancer treatment.

• BMB Reports
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• 제47권6호
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• pp.336-341
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• 2014

Endothelial cell protein C receptor (EPCR) plays important roles in blood coagulation and inflammation. EPCR activity is markedly changed by ectodomain cleavage and release as the soluble EPCR. EPCR can be shed from the cell surface, which is mediated by tumor necrosis factor-${\alpha}$ converting enzyme (TACE). Oroxylin A (OroA), a major component of Scutellaria baicalensis Georgi, is known to exhibit anti-angiogenic, antiinflammation, and anti-invasive activities. However, little is known about the effects of OroA on EPCR shedding. Data showed that OroA induced potent inhibition of phorbol-12-myristate 13-acetate (PMA), tumor necrosis factor (TNF)-${\alpha}$, interleukin (IL)-$1{\beta}$ and on cecal ligation and puncture (CLP)-induced EPCR shedding through suppression of TACE expression and activity. In addition, treatment with OroA resulted in reduced PMA-stimulated phosphorylation of p38, extracellular regulated kinases (ERK) 1/2, and c-Jun N-terminal kinase (JNK). These results demonstrate the potential of OroA as an anti-sEPCR shedding reagent against PMA and CLP-mediated EPCR shedding.

• Journal of Microbiology and Biotechnology
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• 제24권5호
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• pp.719-723
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• 2014

Caspases are a family of cysteine proteases that play an important role in the apoptotic pathway. Caspase-6 is an apoptosis effector that cleaves a variety of cellular substrates. The active form of the enzyme is required for use in research. However, it has been difficult to obtain sufficient quantities of active caspase-6 from Escherichia coli. In the present study, we constructed a caspase-6 with a 23-amino-acid deletion in the pro-domain. This engineered enzyme was expressed as a soluble protein in E. coli and was purified using affinity resin. In vitro enzyme assay and cleavage analysis revealed that the engineered active caspase-6 protein had characteristics similar to those of wild-type caspase-6. This novel method can be a valuable tool for obtaining active caspase-6 that can be used for screening caspase-6-specific substrates, which in turn can be used to elucidate the function of caspase-6 in apoptosis.