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. 1996 Apr 2;93(7):2975–2980. doi: 10.1073/pnas.93.7.2975

Intradimerically tethered DNA topoisomerase II is catalytically active in DNA transport.

J E Lindsley 1
PMCID: PMC39745  PMID: 8610153

Abstract

A covalently cross-linked dimer of yeast DNA topoisomerase II was created by fusing the enzyme with the GCN4 leucine zipper followed by two glycines and a cysteine. Upon oxidation of the chimeric protein, a disulfide bond forms between the two carboxyl termini, covalently and intradimerically cross-linking the two protomers. In addition, all nine of the cysteines naturally occurring in topoisomerase II have been changed to alanines in this construct. This cross-linked, cysteine-less topoisomerase II is catalytically active in DNA duplex passage as indicated by ATP-dependent DNA supercoil relaxation and kinetoplast DNA decatenation assays. However, these experiments do not directly distinguish between a "one-gate" and a "two-gate" mechanism for the enzyme.

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