• 한국미생물·생명공학회지
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• 제37권2호
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• pp.105-109
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• 2009

단백질 인산화를 통한 세포내 신호전달기구 중 serine/threonine kinase에 속하는 Akt/PKB는 세포 생존과 사멸, 당대사 등을 조절하는 것으로 알려져 있다. 이러한 이유로, Akt/PKB 단백질은 천연물질들로부터 항암제를 탐색하기 위한 한 가지 target으로 사용되어 왔다. 본 연구에서는 Akt/PKB 단백질을 대량으로 생산하기 위하여 대장균의 단백질 발현 시스템을 이용하여 human Akt/PKB 단백질을 발현시켰다. 대장균에서 대량 발현된 Akt는 일반적인 조건에서는 inclusion body를 형성하였다. 배양온도 $27^{\circ}C$에서 0.01-0.09 mM IPTG로 발현 유도 시 발현된 human Akt/PKB 단백질 상당 부분이 가용화 되었다. 발현된 Akt kinase를 $Ni^{2+}$-NTA agarose column으로 정제하고, anti-Akt antibody를 이용하여 정제된 단백질이 Akt kinase 임을 확인하였다. 정제된 human Akt/PKB 단백질은 세포추출물에 존재하는 인산화 단백질을 이용하여 in vitro에서 인산화 되었으며, 인산화된 활성형 human Akt/PKB protein kinase는 human Akt/PKB protein kinase 특이 형광 peptide를 특이적으로 인산화하였다.

• 대한두경부종양학회지
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• 제21권2호
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• pp.158-164
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• 2005

Purpose: The serine/threonine kinase Akt was described to inhibit apoptosis in cancer. This study was to examine the effect of Gleevec on head and neck squamous cell carcinoma(HNSCC) through the mechanism of Akt. Experimental Design: Gleevec was introduced into the HNSCC cell lines UMSCC10B, HN12 and HN30 in a range of concentrations. Cell viability was assessed by clonogenic survival analysis. Targets of Gleevec(PDGFR, c-Kit, and c-Abl) were evaluated by Western blot. HNSCC tissue samples were stained for PDGFR, c-Kit and phosphorylated Akt. Akt phosphorylation following Gleevec treatment was assessed using Western blot. Akt siRNA was used to as the positive control. Results: Colony forming efficiency decreased with an increase in concentration of Gleevec. Expressions of PDGFR, c-Kit, and c-Abl were observed in HNSCC cells. Immunohistochemistry confirmed high expression of PDGFR, c-Kit, and p-Akt in human HNSCC tissues. Akt kinase activity was significantly inhibited with increasing concentration of Gleevec in HNSCC cells, and near complete dephosphorylation of Akt was observed at $6{\mu}M$ of Gleevec in the UMSCC10B and HN30 cell lines. Conclusions: Gleevec at clinically comparable concentrations caused a dose dependant decrease in HNSCC survival. The decreased cell survival was related to the inhibition of Akt kinase activity and dephosphorylation of Akt. Akt signaling pathway may be a relevant target for Gleevec in treating HNSCC.

• 약학회지
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• 제48권2호
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• pp.105-110
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• 2004

Akt (or Protein Kinase B; PKB) is a serine/threonine kinase and is activated by phosphoinositide 3-kinase (PI3K) pathway. Recent evidence indicates that the abnormal activities or expression of Akt is closely associated with cancer, diabetes and neuro-degenerative diseases. These findings mean that Akt is likely to be a new therapeutic target for the treatment of disease. Here, we screened the effects of dithiolo-dithione derivatives such as SWU-20004, SWU-20009 and SWU-20025 on Akt activities. Among these compounds, only SWU-20009 (2-Thioxo-[1,3]dithiolo[4,5- $\beta$][1,4]dithiine-5,6-dicarboxylic acid dimethyl ester) inhibited the growth of KATOIII cell at micromolar range of concentration. Further investigation also revealed that SWU-20009 inhibited cellular Akt activity and induced apoptotic cell death.

• BMB Reports
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• 제35권1호
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• pp.106-115
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• 2002

The serine/threonine kinase Akt has been intensely studied for its role in growth factor-mediated cell survival for the past 5 years. On the other hand, the ongoing research effort has recently uncovered novel regulatory mechanisms and downstream effectors of Akt that demonstrate the involvement of Akt in other cellular functions such as cell cycle progression, angiogenesis, and cancer cell invasion/metastasis. Furthermore, recent studies using whole model organisms suggest additional roles for Akt in important diseases such as aging and diabetes. The following review addresses these recent advances in the understanding of Akt function.

• 한국응용과학기술학회지
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• 제25권3호
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• pp.370-379
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• 2008

Akt, a serine/threonine protein kinase as a viral oncogene, is a critical regulator of PI3K-mediated cell proliferation and survival. On translocation, Akt is phosphorylated and activated, ultimately resulting in stimulation of cell growth and survival. As a part of our program toward the novel Akt1 inhibitors with potent activity over PI3K signaling pathway, we found primary hit compound 2 with an $IC_{50}$ value of $620\mu}M$ from protein kinase focused library. Based on the structural features of 2, new 1,3,4-thiadiazole derivatives were designed by the introduction of aromatic and heteroaromatic moieties onto thiadiazole nucleus. In this work, a series of 1,3,4-thiadiazole derivatives 1a-1 were synthesized and evaluated for Akt1 inhibitory activity.

• 한국미생물·생명공학회지
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• 제30권3호
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• pp.235-240
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• 2002

방선균은 토양속에 서식하는 그람 양성 세균으로 세포성 장의 어느 시기에 영양세포가 이어져 연쇄상의 기균사를 형성하고 그 끝에 포자를 형성하는 동시에 생리학적 분화로 표현되는 다양한 이차대사물질을 생산한다. 이들의 복잡한 생활사에 따른 분화에는 진핵생물의 ser/thr protein kinase와 원핵생물의 his/asp acid protein kinase 등과 같은 다양한 신호전달 단백질들이 조절을 담당하고 있다. Akt kinase는 진핵생물에서 보고된 ser/thr kinase로.세포내의 다양한 신호전달기구를 조절하고 있으며, 세포내의 Akt kinase의 활성화 또는 불활성화가 세포 증식, 분화, 생존, 세포사등의 신호전달에 결정적인 역할을 담당한다. 방선균으로부터 Akt kinase와 유사한 기능을 갖는 신호전달 단백질을 규명하기 위하여, Akt kinae의 target단백질들의 인산화 부위 보존영역으로부터 나타나는 아미노산의 consensus sequence를 기초로 하여 형광물질로 라벨시킨 합성 peptide(FITC-TRRSRfESIT)를 제작하였다 제작한 기질 peptide에 인산화가 일어나면 아가 로스 전기영동상에서의 운동성에 차이가 나타나고, 이를 자외선하에서 형광 peptide를 관찰하는 방법으로 인산화 assay를 실시하였다. S. griseus IFO 13350을 배양한 cell-free extract로부터 ammonium sulfate fractionation과 DEAE-Sepharose, Mono Q, Resource Phenyl-Superose, Gel permeation 등 수 단계의 column chromatography를 통하여 Akt 유사 단백질을 정제하였다. 그 결과 방선균에도 고등생 물의 Akt와 유사한 기질특이성을 갖는 인산화 단백질이 존재하는 것으로 판단되었으며, 그 중의 하나는 분자량이 39 kDa 정도의 크기를 갖는 단백질로 판명되었다. 지금까지의 인산화 단백질 연구는 활성측정법이 어려워 연구자들에게 많은 제한을 주어 왔지만, 본 연구에서 사용한 합성 peptide를 이용하는 방법을 보다 다양한 인산화 단백질에 대하여 적용한다면, 인산화 단백질 및 조절물질 개발에 많은 도움이 될 수 있을 것으로 예상된다.

• 대한약학회:학술대회논문집
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• 대한약학회 2003년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.2-2
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• pp.168.2-168.2
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• 2003

Akt/Ptotein Kinase B (PKB) is a serine/threonine kinase and activated by PI3K pathway. Akt/PKB regulates a variety of cellular responses including proliferations, differentiations and insulin signaling pathway. Recent evidence also indicates that the abnormal activities or expression of Akt/PKB is closely associated with cancer, diabetes and neuro-degenerative diseases. These findings mean that Akt/PKB is likely to be a new therapeutic target for the treatment of disease. (omitted)

• Biomolecules & Therapeutics
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• 제31권1호
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• pp.127-138
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• 2023

Glycogen synthase kinase-3β (GSK-3β) is an important serine/threonine kinase that implicates in multiple cellular processes and links with the neurodegenerative diseases including Alzheimer's disease (AD). In this study, structure-based virtual screening was performed to search database for compounds targeting GSK-3β from Enamine's screening collection. Of the top-ranked compounds, 7 primary hits underwent a luminescent kinase assay and a cell assay using human neuroblastoma SH-SY5Y cells expressing Tau repeat domain (Tau_RD) with pro-aggregant mutation ΔK280. In the kinase assay for these 7 compounds, residual GSK-3β activities ranged from 36.1% to 90.0% were detected at the IC₅₀ of SB-216763. In the cell assay, only compounds VB-030 and VB-037 reduced Tau aggregation in SH-SY5Y cells expressing ΔK280 Tau_RD-DsRed folding reporter. In SH-SY5Y cells expressing ΔK280 Tau_RD, neither VB-030 nor VB-037 increased expression of GSK-3α Ser21 or GSK-3β Ser9. Among extracellular signal-regulated kinase (ERK), AKT serine/threonine kinase 1 (AKT), mitogen-activated protein kinase 14 (P38) and mitogenactivated protein kinase 8 (JNK) which modulate Tau phosphorylation, VB-037 attenuated active phosphorylation of P38 Thr180/ Tyr182, whereas VB-030 had no effect on the phosphorylation status of ERK, AKT, P38 or JNK. However, both VB-030 and VB-037 reduced endogenous Tau phosphorylation at Ser202, Thr231, Ser396 and Ser404 in neuronally differentiated SH-SY5Y expressing ΔK280 Tau_RD. In addition, VB-030 and VB-037 further improved neuronal survival and/or neurite length and branch in mouse hippocampal primary culture under Tau cytotoxicity. Overall, through inhibiting GSK-3β kinase activity and/or p-P38 (Thr180/Tyr182), both compounds may serve as promising candidates to reduce Tau aggregation/cytotoxicity for AD treatment.

• BMB Reports
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• 제57권6호
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• pp.305-310
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• 2024

T-plastin (PLST), a member of the actin-bundling protein family, plays crucial roles in cytoskeletal structure, regulation, and motility. Studies have shown that the plastin family is associated with the malignant characteristics of cancer, such as circulating tumor cells and metastasis, by inducing epithelial-mesenchymal transition (EMT) in various cancer cells. However, the role of PLST in the EMT of human lung cancer cells remains unclear. In this study, we observed that PLST overexpression enhanced cell migratory and invasive abilities, whereas its downregulation resulted in their suppression. Moreover, PLST expression levels were associated with the expression patterns of EMT markers, including E-cadherin, vimentin, and Slug. Furthermore, the phosphorylation levels of focal adhesion kinase (FAK) and AKT serine/threonine kinase (AKT) were dependent on PLST expression levels. These findings indicate that PLST induces the migration and invasion of human lung cancer cells by promoting Slug-mediated EMT via the FAK/AKT signaling pathway.

• Journal of Microbiology and Biotechnology
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• 제18권2호
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• pp.377-382
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• 2008

Interleukin-2 plays a significant role in T cell proliferation. Here, we report the role of IL-2 in angiogenesis. IL-2 increased the ROS level and phosphorylation of Akt in human umbilical vein endothelial cells (HUVECs). IL-2 increased angiogenesis in an animal model and tube formation in HUVECs. The effect of IL-2 on angiogenesis and tube formation was mediated by ROS and Akt. This is the first report that IL-2 promotes angiogenesis.