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DNA gyrase is a bacterial type II topisomerase 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 from the over producing strains JMtacA and JMtacB1 and is supplied as an A2B2 complex.
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 enzyme and linearised pBR322 marker.
Cleavage specific enzyme available separately on request.
These kits are designed for assaying cell extracts containing E. coli gyrase and partially purified fractions and contain relaxed DNA substrate, Assay buffer, Dilution buffer and control supercoiled DNA.
These kits can be used to test the effects of potential ATPase inhibitors. For example, the coumarin drugs such as novobiocin inhibit the action of DNA gyrase by competitively inhibiting the hydrolysis of ATP thus preventing supercoiling.
These assays are microtitre plate-based and thus large numbers of compounds can be screened in a relatively short period of time. They also continuous assays which can provide more information than an end point assay.
The kit is supplied with sufficient E. coli gyrase enzyme, plasmid DNA substrate, buffers and other assay components for 100 assays. The enzyme is supplied at a concentration of 5 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)