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During DNA replication, DNA helicase (DnaB) unwinds DNA using energy from ATP hydrolysis. When the helicase ATPase is blocked, DNA unwinding stops too, so this ATPase reaction serves as a drug discovery screen for helicase inhibitors.
Pack Sizes
Both sizes include the E. coli DNA helicase. The only difference is the number of assays: DNAB100KE provides reagents for 100 assays and DNAB500KE for 500 assays (384-well format), with component volumes scaled accordingly.
| 100 assays (DNAB100KE) | 100 assays; enzyme included |
|---|---|
| 500 assays (DNAB500KE) | 500 assays; enzyme included |
Kit Components
| Component | 100 assays (DNAB100KE) | 500 assays (DNAB500KE) |
|---|---|---|
| 10 x Assay buffer | 500 µl | 2 ml |
| 100 x DNA | 35 µl | 170 µl |
| 100 x E. coli DNA helicase | 30 µl | 150 µl |
| 100 x ATP | 35 µl | 170 µl |
| Phosphate detection dye (also sold separately as MicroMolar Phosphate Assay Reagent, BHT16300173) | 5 ml | 25 ml |
Final reaction conditions: 20 mM HEPES pH 7.5, 20 mM potassium glutamate, 1 mM DTT, 0.005% Triton X-100, 10 mM MgCl2, 250 nM DNA, 0.25 mM ATP and 200 nM DNA helicase.
Assay Principle
The kit detects the phosphate produced when E. coli DNA helicase hydrolyzes ATP in the presence of DNA. A phosphate detection dye is added and absorbance is read at 650 nm; inhibitors of the helicase ATPase lower the signal.
Specifications
| Assay Target | E. coli DNA helicase ATPase |
|---|---|
| Target Type | Helicase |
| Organism | E. coli |
| Assay Type | Enzyme Activity Assay; Inhibitor Screening Assay |
| Detection Method | Colorimetric |
| Detection Wavelength | Absorbance 650 nm |
| Plate Format | 384-well plate |
| Instrument Required | Absorbance microplate reader |
| Assay Time | About 65 min (60 min reaction + 5 min detection) |
| Publication | Nakano T. et al., J. Biol. Chem. 288: 4649-4658 (2013) |
| Sizes | 100 assays, 500 assays |
| Storage | Please contact us to confirm storage conditions. |
| Shipping | Shipped frozen. |
Protocol
The manufacturer's protocol describes the procedure and reaction volumes.
- E. coli DNA Helicase ATPase assay (PDF, see Documents): Covers catalog # DNAB100KE, DNAB500KE.
Safety & Handling
Can’t find the exact format you need? We can help you source or customize this E. coli DNA helicase ATPase assay — including larger pack sizes, bulk reagents, bulk enzyme, or a protocol adapted to your samples. Click Talk to a Scientist, email support@biohippo.com, or explore our Research Services. Our team will be in touch shortly.