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BioAssay: AID 652256

qHTS Assay for Inhibitors of Mammalian Selenoprotein Thioredoxin Reductase 1 (TrxR1): DTNB Assay

The selenoprotein thioredoxin reductase (TrxR; EC 1.8.1.9) is a FAD containing homodimeric pyridine nucleotide-disulfide oxidoreductase with many cellular roles. Together with NADPH and its prime substrate thioredoxin (Trx), the enzyme forms the core of the Trx system. The mammalian Trx system exerts a wide spectrum of functions including redox regulation, antioxidant defense, regulation of more ..
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 Tested Compounds
 Tested Compounds
All(54)
 
 
Active(49)
 
 
Inactive(5)
 
 
 Tested Substances
 Tested Substances
All(54)
 
 
Active(49)
 
 
Inactive(5)
 
 
AID: 652256
Data Source: NCGC (TrxR202)
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2013-04-02

Data Table ( Complete ):           Active    All
Target
BioActive Compounds: 49
Depositor Specified Assays
AIDNameTypeComment
488771Probe Development Summary for Modulators of Mammalian Selenoprotein Thioredoxin Reductase 1 (TrxR1)summary
Description:
The selenoprotein thioredoxin reductase (TrxR; EC 1.8.1.9) is a FAD containing homodimeric pyridine nucleotide-disulfide oxidoreductase with many cellular roles. Together with NADPH and its prime substrate thioredoxin (Trx), the enzyme forms the core of the Trx system. The mammalian Trx system exerts a wide spectrum of functions including redox regulation, antioxidant defense, regulation of transcription factors as well as support of cell growth and replication. Many of these functions involve the reduction of Trx, which may subsequently reduce a number of different substrates including ribonucleotide reductase, peroxiredoxins or NFkB. Mammalian TrxR itself also has a broad substrate specificity, reducing both protein and non-protein substrates, including low molecular weight compounds such as dehydroascorbate, lipoic acid, ubiquinone, and juglone. In addition, several drugs in clinical use for anticancer treatment are indeed known to target TrxR1.

This orthogonal DTNB assay seeks to further characterize cherry picks.

NIH Molecular Libraries Probe Production Network [MLPCN]
NIH Chemical Genomics Center [NCGC]

MLPCN Grant: MH090846
Assay Provider: Elias Arner, Karolinska Institute
Protocol
15 nM TrxR1 is reduced by 250 microM NADPH in TE buffer [50 mM Tris-HCl (pH 7.5), 2 mM EDTA] in a 96-well microtiter plate. Compounds are then added and incubated for 30 min at 30 degrees C. During compound incubation, NADPH consumption is followed by the decrease in absorbance at 340 nm using a VersaMax microplate reader (Molecular Devices) to determine substrate potential of the compound. DTNB is subsequently added to a final concentration of 2.5 mM, and the formation of TNB- is followed at 412 nm at 30 degrees C to assess inhibitory activity of the compound.
Comment
Compounds were an IC50 was determined < 100 uM were considered "active" and assigned a score of 90; compounds were the IC50 >= 100 uM or could not be determined were considered "inactive" and assigned a score of 10.
Result Definitions
TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1DTNB 412 IC50 uMFloatμM
Additional Information
Grant Number: MH090846

Data Table (Concise)
Classification
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