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. 1989 Aug;86(16):6121–6125. doi: 10.1073/pnas.86.16.6121

Template supercoiling during ATP-dependent DNA helix tracking: studies with simian virus 40 large tumor antigen.

L Yang 1, C B Jessee 1, K Lau 1, H Zhang 1, L F Liu 1
PMCID: PMC297787  PMID: 2548199

Abstract

Incubation of topologically relaxed plasmid DNA with simian virus 40 (SV40) large tumor antigen (T antigen), ATP, and eubacterial DNA topoisomerase I resulted in the formation of highly positively supercoiled DNA. Eukaryotic DNA topoisomerase I could not substitute for eubacterial DNA topoisomerase 1 in this reaction. Furthermore, the addition of eukaryotic topoisomerase I to a preincubated reaction mixture containing both T antigen and eubacterial topoisomerase I caused rapid relaxation of the positively supercoiled DNA. These results suggest that SV40 T antigen can introduce topoisomerase-relaxable supercoils into DNA in a reaction coupled to ATP hydrolysis. We interpret the observed T antigen supercoiling reaction in terms of a recently proposed twin-supercoiled-domain model that describes the mechanics of DNA helix-tracking processes. According to this model positive and negative supercoils are generated ahead of and behind the moving SV40 T antigen, respectively. The preferential relaxation of negative supercoils by eubacterial DNA topoisomerase I explains the accumulation of positive supercoils in the DNA template. The supercoiling assay using DNA conformation-specific eubacterial DNA topoisomerase I may be of general use for the detection of ATP-dependent DNA helix-tracking proteins.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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