E. coli Topoisomerase IV

Topo IV (from Escherichia coli) is prepared by overexpressing the parC and parE subunits in E. coli and purifying them by methods adapted from Peng and Marians, 1999¹.

The parC and parE subunits are purified to >95% purity as judged by SDS-PAGE. The topo IV is supplied as a heterotetramer complex in Dilution buffer.

It is recommended that the enzyme is aliquoted to avoid repeated freeze-thaw cycles. Store at -80ºC.

All enzyme is supplied with 5X concentrated Assay Buffer and Dilution buffers which are also available separately.

See technical documents below for more detailed information and lot specific activities.

Technical Documents

Cat No.	Product	Price	Quantity
T4001	E. coli Topo IV100 U	£133
T4002	E. coli Topo IV500 U	£499
T4003	E. coli Topo IV1000 U	£995
T4004	E. coli Topo IV2000 U	£1,875

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These contain the enzyme and the supercoiled DNA substrate in addition to the Assay and Dilution buffers for relaxation reactions. 1 U of topo IV will relax 0.5 µg supercoiled pBR322 DNA in 30 minutes at 37°C. 

Technical Documents

Cat No.	Product	Price	Quantity
R4001	E. coli Topo IV Relaxation Kit 1100 U of E. coli Topo IV and 50 µg of Supercoiled DNA	£220
R4002	E. coli Topo IV Relaxation Kit 2500 U of E. coli Topo IV and 50 µg of Supercoiled DNA	£810
R4003	E. coli Topo IV Relaxation Kit 31000 U of E. coli Topo IV and 50 µg of Supercoiled DNA	£1,603
R4004	E. coli Topo IV Relaxation Kit 42000 U of E. coli Topo IV and 50 µg of Supercoiled DNA	£3,016

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These contain the enzyme and the catenated kDNA substrate in addition to the Assay and Dilution buffers for decatenation reactions. 1 U of topo IV will decatenate 200 ng of kDNA when incubated in 1X Assay buffer in a total reaction volume of 30 µl at 37°C for 30 minutes. 

Technical Documents

Cat No.	Product	Price	Quantity
D4001	E. coli Topo IV Decatenation Kit 1100 U of E. coli Topo IV and 20 µg kDNA	£220
D4002	E. coli Topo IV Decatenation Kit 2500 U of E. coli Topo IV and 100 µg kDNA	£810
D4003	E. coli Topo IV Decatenation Kit 31000 U of E. coli Topo IV and 200 µg kDNA	£1,603
D4004	E. coli Topo IV Decatenation Kit 42000 U of E. coli Topo IV and 400 µg kDNA	£3,016

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These kits are designed for assaying cell extracts and partially purified fractions containing over-expressed topo IV and contain supercoiled DNA substrate, Assay buffer, Dilution buffer, control relaxed DNA and stop buffer/loading dye.

Technical Documents

Cat No.	Product	Price	Quantity
TCE001	E. coli Topo IV Assay Kit 1 for Cell ExtractsFor 100 assays	£136
TCE002	E. coli Topo IV Assay Kit 2 for Cell ExtractsFor 500 assays	£402
TCE003	E. coli Topo IV Assay Kit 3 for Cell ExtractsFor 1000 assays	£799
TCE004	E. coli Topo IV Assay Kit 4 for Cell ExtractsFor 2000 assays	£1,527

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These kits are designed specifically for cleavage reactions. They contain topo IV enzyme, 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.

Technical Documents

Cat No.	Product	Price	Quantity
TCK001	E. coli Topo IV Cleavage Kit 1100 U of E. coli Topo IV and 50 µg of supercoiled DNA	£399

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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 topoisomerase IV 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.

Technical Documents

Cat No.	Product	Price	Quantity
ATPECT001	Kit for 100 ATPase assaysFor determination of DNA-dependent ATPase activity; contains E. coli topoIV, buffers, ATP, PK-LDH, PEP, NADH and linear pBR322 DNA substrate.	£1,472

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The kit is supplied with sufficient E. coli topo IV 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)

Technical Documents

Cat No.	Product	Price	Quantity
TRIV01	HTS Assay Kit - E. coli Topo IV 1-5 KitsFor 100 assays per plate	£440
TRIV02	HTS Assay Kit - E. coli Topo IV 6+ KitsFor 100 assays per plate	£346

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