HTS for inhibitor screening and evaluation of glucose-6-phosphate dehydrogenase inhibitors.
- Non-radioactive assay.
- Homogenous “mix-incubate-measure” type assay. Can be readily automated on HTS liquid handling system.
- Rapid and reliable. Can be completed in 30 minutes and no 37°C heater is needed.
- Method: OD565nm
- Samples: Glucose-6-phosphate dehydrogenase
GLUCOSE-6-PHOSPHATE DEHYDROGENASE (G6PDH) is a cytosolic enzyme in the pentose phosphate pathway which supplies reducing energy to cells by maintaining the level of the co-enzyme nicotinamide adenine dinucleotide phosphate (NADPH). G6PDH reduces nicotinamide adenine dinucleotide phosphate (NADP) to NADPH while oxidizing glucose-6-phosphate (G6P). Humans with a genetic deficiency of G6PDH are predisposed to non-immune hemolytic anemia. Recently, studies have found G6PDH plays a critical role in survival, proliferation, and metastasis of cancer cells. Therefore, inhibitors of the enzyme are attractive candidates for new cancer therapeutics. BioAssay Systems non-radioactive, colorimetric G6PDH assay is based on the reduction of the tetrazolium salt MTT in a NADPH-coupled enzymatic reaction to a reduced form of MTT which exhibits an absorption maximum at 565 nm. The increase in absorbance at 565 nm is proportional to the enzyme activity. The percent inhibition of a test compound can be determined by comparing the activity of G6PDH treated with a test compound to the activity of untreated G6PDH.