scholarly journals A Novel Assay Method to Determine the β-Elimination of Se-Methylselenocysteine to Monomethylselenol by Kynurenine Aminotransferase 1

Antioxidants ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 139
Author(s):  
Selvam ◽  
Björnstedt
Keyword(s):  
Cancer Cells ◽  
Thioredoxin Reductase ◽  
Accurate Method ◽  
Spectrophotometric Assay ◽  
Assay Method ◽  
Mechanistic Studies ◽  
Activity Assay ◽  
Kynurenine Aminotransferase ◽  
Selenium Compound ◽  
Beta Elimination

Kynurenine aminotransferase 1 (KYAT1 or CCBL1) plays a major role in Se-methylselenocysteine (MSC) metabolism. It is a bi-functional enzyme that catalyzes transamination and beta-elimination activity with a single substrate. KYAT1 produces methylselenol (CH3SeH) via β-elimination activities with MSC as a substrate. This methylated selenium compound is a major cytotoxic selenium metabolite, causing apoptosis in a wide variety of cancer cells. Methylselenol is volatile and possesses extraordinary nucleophilic properties. We herein describe a simple spectrophotometric assay by combining KYAT1 and thioredoxin reductase (TrxR) to detect CH3SeH in a coupled activity assay. The metabolite methylselenol and its oxidized form from MSC metabolism is utilized as a substrate for TrxR1 and this can be monitored spectroscopically at 340 nm. Our results show the feasibility of monitoring the β-elimination of KYAT1 by our assay and the results were compared to the previously described β-elimination assays measuring pyruvate. By using known inhibitors of KYAT1 and TrxR1, we further validated the respective reaction. Our data provide a simple but accurate method to determine the β-elimination activity of KYAT1, which is of importance for mechanistic studies of a highly interesting selenium compound.

Pyrrolizines as Potential Anticancer Agents: Design, Synthesis, Caspase-3 activation and Micronucleus (MN) Induction

2019 ◽  
Vol 18 (15) ◽  
pp. 2124-2130
Author(s):  
Amany Belal
Keyword(s):  
Cancer Cells ◽  
Cell Lines ◽  
Anticancer Agents ◽  
Caspase 3 ◽  
Assay Method ◽  
Breast Adenocarcinoma ◽  
Compound I ◽  
Design Synthesis ◽  
Ic50 Values ◽  
New Compounds

Background: For further exploration of the promising pyrrolizine scaffold and in continuation of our previous work, that proved the potential anticancer activity of the hit compound I, a new series of pyrrolizines 2-5 and 7-9 were designed and synthesized. Methods: Structures of the new compounds were confirmed by IR, 1H-NMR, 13C-NMR and elemental analysis. Antitumor activity for the prepared compounds against human breast adenocarcinoma (MCF-7), liver (HEPG2) and colon (HCT116) cancer cell lines was evaluated using SRB assay method. Result: Compounds 2, 3 and 5 were the most potent on colon cancer cells, their IC50 values were less than 5 µM. Compounds 2, 3 and 8 were the most potent on liver cancer cells, their IC50 values were less than 10 µM. As for MCF7, compounds 2, 7, 8 and 9 were the most active with IC50 values less than 10 µM. We can conclude that combining pyrrolizine scaffold with urea gave abroad spectrum anticancer agent 2 against the three tested cell lines. Micronucleus assays showed that compounds 2, 3, 8 are mutagenic and can induce apoptosis. In addition, caspase-3 activation was evaluated and compound 2 showed increase in the level of caspase-3 (9 folds) followed by 3 (8.28 folds) then 8 (7.89 folds). Conclusion: The obtained results encourage considering these three compounds as novel anticancer prototypes.

scholarly journals ADCK1 activates the β-catenin/TCF signaling pathway to promote the growth and migration of colon cancer cells

2021 ◽  
Vol 12 (4) ◽  
Author(s):  
Yong Ji ◽  
Yiqian Liu ◽  
Changchun Sun ◽  
Lijiang Yu ◽  
Zhao Wang ◽  
...  
Keyword(s):  
Colon Cancer ◽  
Cancer Cells ◽  
Signaling Pathway ◽  
Colony Formation ◽  
Activation Mechanism ◽  
Colon Cancer Cells ◽  
Mechanistic Studies ◽  
T Cell Factor ◽  
And Migration

AbstractAs a result of mutations in the upstream components of the Wnt/β-catenin signaling pathway, this cascade is abnormally activated in colon cancer. Hence, identifying the activation mechanism of this pathway is an urgent need for the treatment of colon cancer. Here, we found an increase in ADCK1 (AarF domain-containing kinase 1) expression in clinical specimens of colon cancer and animal models. Upregulation of ADCK1 expression promoted the colony formation and infiltration of cancer cells. Downregulation of ADCK1 expression inhibited the colony formation and infiltration of cancer cells, in vivo tumorigenesis, migration, and organoid formation. Molecular mechanistic studies demonstrated that ADCK1 interacted with TCF4 (T-cell factor 4) to activate the β-catenin/TCF signaling pathway. In conclusion, our research revealed the functions of ADCK1 in the development of colon cancer and provided potential therapeutic targets.

scholarly journals EF24 induces ROS-mediated apoptosis via targeting thioredoxin reductase 1 in gastric cancer cells

Oncotarget ◽  
2016 ◽  
Vol 7 (14) ◽  
pp. 18050-18064 ◽  
Author(s):  
Peng Zou ◽  
Yiqun Xia ◽  
Weiqian Chen ◽  
Xi Chen ◽  
Shilong Ying ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cancer Cells ◽  
Thioredoxin Reductase ◽  
Gastric Cancer Cells ◽  
Thioredoxin Reductase 1

scholarly journals A sensitive, accurate, and high-throughput gluco-oligosaccharide oxidase based HRP colorimetric method for lytic polysaccharide monooxygenase activity assay

2021 ◽  
Author(s):  
Gang Liu
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Direct Method ◽  
Total Concentration ◽  
Colorimetric Method ◽  
Industrial Applications ◽  
Assay Method ◽  
Activity Assay ◽  
Lytic Polysaccharide Monooxygenase ◽  
Monooxygenase Activity

Abstract Background The AA9 (auxiliary activities) family of lytic polysaccharide monooxygenases (AA9 LPMOs) are ubiquitous and diverse group of enzymes amongst the fungal kingdom. They catalyze the oxidative cleavage of glycosidic bonds in lignocellulose and exhibit great potential for secondary biorefinery applications. Screening of AA9 LPMOs for desirable properties is crucial for biorefinery industrial applications. However, robust, high-throughput and direct method for AA9 LPMO activity assay, which is prerequisite for screening of LPMOs with excellent properties, is still lacking. Here, we have described a gluco-oligosaccharide oxidase (GOOX) based horseradish peroxidase (HRP) colorimetric method for AA9 LPMO activity assay. Results We cloned and expressed a GOOX gene from Sarocladium strictum in Trichoderma reesei, purified the recombinant SsGOOX, validated its properties, and set up a SsGOOX based HRP colorimetric method for cellobiose concentration assay. Then we expressed two AA9 LPMOs from Thielavia terrestris, TtAA9F and TtAA9G in T. reesei, purified the recombinant proteins, and analyzed their product profiles and regioselectivity towards phosphoric acid swollen cellulose (PASC). TtAA9F was characterized as a C1 type (class 1) LPMO, while TtAA9G was characterized as a C4 type (class 2) LPMO. Finally, the SsGOOX based HRP colorimetric method was used to quantify the total concentration of reducing lytic products from LPMO reaction, and consequently, the activities of both C1 and C4 types of LPMOs were analyzed. These LPMOs could be effectively analyzed with limits of detection (LoDs) lower than 30 nmol/L, and standard curves between A515 and LPMO concentrations with determination coefficients greater than 0.994 were obtained. Conclusions A novel, sensitive and accurate assay method that directly targets the main activity of both C1 and C4 type of AA9 LPMOs was established. This method is easy to use and could be performed on a microtiter plate ready for high-throughput screening of AA9 LPMOs with high properties.

Thioredoxin reductase inhibition by new trans-platinum(II) complexes elicit cytotoxicity in cisplatin resistant ovarian cancer cells

2021 ◽  
Vol 177 ◽  
pp. S106-S107
Author(s):  
Valeria Scalcon ◽  
Mariafrancesca Hyeraci ◽  
Alessandra Folda ◽  
Luca Labella ◽  
Fabio Marchetti ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Cancer Cells ◽  
Thioredoxin Reductase ◽  
Ovarian Cancer Cells

Ruthenium polypyridyl complexes as inducer of ROS-mediated apoptosis in cancer cells by targeting thioredoxin reductase

Metallomics ◽  
2014 ◽  
Vol 6 (8) ◽  
pp. 1480-1490 ◽  
Author(s):  
Zuandi Luo ◽  
Lianling Yu ◽  
Fang Yang ◽  
Zhennan Zhao ◽  
Bo Yu ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Cell Apoptosis ◽  
Thioredoxin Reductase ◽  
Cancer Growth ◽  
Ros Generation ◽  
Ruthenium Polypyridyl ◽  
Polypyridyl Complexes ◽  
Intracellular Ros ◽  
Ruthenium Polypyridyl Complexes ◽  
Apoptosis In Cancer Cells

Ruthenium polypyridyl complexes inhibit cancer growth by targeting TrxR and promote the intracellular ROS generation, ultimately triggering mitochondria-mediated cell apoptosis.

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