• Microbiology and Biotechnology Letters
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• 제37권2호
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• pp.105-109
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• 2009

Among signal transduction systems by protein phosphorylation Akt/PKB protein kinase which is one of serine/threonine kinases, is known to regulate the survival and death of the cell and glucose metabolism. Thus, Akt/PKB protein kinase has been used as one of the target proteins to find anti-cancer agents from natural products. In this study, human Akt/PKB protein kinase was expressed in Escherichia coli expression system for the mass production. Human Akt/PKB protein kinase expressed in E. coli formed inclusion body under the general condition. However, most of the expressed protein was solubilized under the culture temperature at $27^{\circ}C$ and 0.01-0.09 mM of IPTG for induction of the protein expression. The expressed protein was purified using $Ni^{2+}$-NTA agarose column and confirmed by using anti-Akt antibody. Subsequently, the purified human Akt/PKB protein kinase was activated by in vitro phosphorylation using cellular extract containing kinases. The activated protein was confirmed to phosphorylate the specific fluorescent peptide specially designed as the artificial substrate for Akt/PKB protein kinase.

• Korean Journal of Head & Neck Oncology
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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.

• YAKHAK HOEJI
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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.

• Journal of the Korean Applied Science and Technology
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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.

• Microbiology and Biotechnology Letters
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• 제30권3호
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• pp.235-240
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• 2002

Streptomycetes is a group of Gram-positive soil bacteria that growas a branching vegetative mycelium leading to the formation of spores, and display a physiological differenti-ation related to the synthesis of many secondary metabolites including antibiotics. Their complex life cycle and multicellular differentiation require various levels of regulation and types of signal transduction systems including eukaryotic-type serine/threonine protein kinases and prokaryotic-type histidine/aspartic acid protein kinases. Akt kinase that was found in cells is a sorine/threonine kinase controlling signal pathway for multi-tude of important cellular events. The activation or inactivation of Akt kinase in the cell is one of the critical regulatory points to deliver cell proliferation, differentiation, survival or apoptosis signal. To find the regula-tory protein homologous to Akt in Streptomyces, the fluorescien-labeled synthetic peptide (FITC-TRRSR-TESIT) was designed from the consensus sequence of target proteins for Akt kinase. From the difference of the mobility between the nonphosphorylated and phosphorylated synthetic peptides on Agarose gel electro-phoresis, the Akt-phosphorylating activity was monitored. The cell-free extract prepared from Streptomyces griseus IFO 13350 and the Akt homologous protein was purified by ammonium sulfate fractionation and many steps of column chromatographies such as, DEAE-Sepharose, Mono Q, Resource Phenyl-Soporose and Gel permeation column chromatographies. As a result, the protein phosphorylating the fluorescien-labeled Akt substrate was identified and it's molecular weight was estimated as 39 kDa on SDS-PAGE.

• Proceedings of the PSK Conference
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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.