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Staphylococcus aureus gyrase is a bacterial type II topoisomerase which can introduce negative supercoils into DNA. It is a target for both quinolone and coumarin drugs and can be used for screening potential antibacterial compounds. It is prepared by overexpressing the subunits in E. coli and then purified.
It is supplied as an A2B2 complex in Dilution Buffer. Store at -80 °C.
All enzyme is supplied with 5X concentrated Assay Buffer and Dilution Buffer which are also available separately. 1 U of gyrase will supercoil 0.5 µg relaxed pBR322 DNA in 30 minutes at 37 °C.
See technical documents below for more detailed information and lot specific activities.
Kits are available containing everything required to perform supercoiling reactions and to test inhibitors. They contain the enzyme, relaxed DNA substrate and the Assay and Dilution buffers.
Some drugs interrupt the DNA breakage-reunion step of the gyrase reaction. This leads to cell death and it is the mechanism behind the action of the quinolones such as nalidixic acid and ciprofloxacin. Cleavage assays are particularly useful in determining if a potential drug acts by this mechanism.
These kits are designed specifically for cleavage reactions. They contain the supercoiled pBR322 DNA substrate and the Assay and Dilution buffers required for DNA cleavage reactions in addition to the S. aureus gyrase and linearised pBR322 marker.
Cleavage specific enzyme available separately on request.
These kits are designed for assaying cell extracts containing gyrase and partially purified fractions and contain relaxed DNA substrate, Assay buffer, Dilution buffer and control supercoiled DNA.
The coumarin drugs inhibit the action of gyrase by competitively inhibiting the hydrolysis of ATP thus preventing supercoiling. These kits can be used to test the effects of potential ATPase inhibitors.
These kits are advantageous because the assays are microtitre plate-based and thus large numbers of compounds can be screened in a relatively short period of time.
The kit is supplied with sufficient S. aureus gyrase enzyme, plasmid DNA substrate, buffers and other assay components* for 100 assays. The enzyme is supplied at a concentration of 10 U/μl in Dilution Buffer. The kit is also supplied with sufficient wash buffers for one 96-well plate. These buffers are supplied as 20X concentrates and must be diluted with ultra pure water prior to use.
More information about this assay can be found on the 'Services' page under 'High/Medium Throughput Assay'.
Kit issued with limited licence for individual use only.
Patent held by Inspiralis Ltd., Norwich, Norfolk, UK. (Patent No. GB0424953.8, US7838230)