We offer a number of topoisomerase assays which can identify inhibitors, determine IC50values or investigate the mechanism of action. They include a medium/high-throughput plate-based assay, gel-based supercoiling, relaxation and decatenation reactions, ATPase assays, cleavage assays and DNA unwinding assays.
Any number or combination of assays can be carried out and there is no minimum number of compounds required.
All assays are performed in-house by experienced researchers. Results are provided as a full written report containing original gel images, gel data and subsequent statistical analysis. We have many years of experience working with these types of assays and can quickly identify positive inhibitors saving you time and money.
Just send us your compounds and we will do the rest.
Hydrolysis of ATP is essential for the activity of topoisomerase II enzymes and inhibition of the ATPase activity is the mechanism by which compounds such as coumarins (e.g. novobiocin) inhibit the enzyme. The activity of the complete enzyme is stimulated by binding DNA (the E.coli GyrB43 domain is not stimulated). The reaction can be followed by a number of methods. In the linked assay, the hydrolysis of ATP by topoisomerase is linked to the conversion of NADH to NAD+ which can then be measured by a change in absorbance at 340nm. This is a continuous assay which allows the course of the reaction to be monitored. Details of the method can be found in the protocols. An alternative assay which uses Malachite Green to directly measure the phosphate produced by the hydrolysis of ATP to ADP is an end-point assay. We are also able to supply this assay if you prefer. Please enquire for more information about this.
Initial set up costs are £500 per assay. Screening costs from £100 per compound for fixed concentration inhibition to £250 per compound for full IC50 determination. However, discounts may be available if larger numbers of assays are performed.