Type II topoisomerase substrate geometry revealed through combined experiment and computation.
- 1National Institutes of Health
Abstract
Type II topoisomerases (topo IIs) are essential enzymes that regulate DNA topology through a strand-passage mechanism in which a duplex DNA (transfer-segment) is passed through a transiently cleaved second duplex DNA (gate-segment). Biochemical and structural approaches have revealed critical details of the binding and cleavage of the gate-segment DNA. However, capture of the transfer-segment DNA has proven more difficult to resolve due to the transient nature of the interaction. Nonetheless, selection of the transfer segment with a specific conformation or orientation relative to the gate segment is predicted to govern aspects of topo II activity, including chiral discrimination and the ability to reduce topological complexity below equilibrium. To determine the conformation and orientation of the transfer-segment relative to the gate segment, we combined experimental single-molecule measurements of topo II unlinking a single DNA crossing with Brownian dynamics simulations of the DNA crossing. By correlating the unlinking rate with the geometric features of the DNA crossing, we obtain the complete three-dimensional preferred crossing geometry for strand passage. Strikingly, the preferred crossing geometries for Escherichia coli topoisomerase IV and Methanosarcina mazei topoisomerase VI are distinct and provide structural models of the DNA synapse selected for strand passage along with a mechanistic basis for their differing activities and biological functions. The approach we develop is generalizable, providing unique insights into the kinetic selection of DNA synapse structure.
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BibTeX
@misc{prajapati2026type,
title = {{Type II topoisomerase substrate geometry revealed through combined experiment and computation.}},
author = {M. N. Prajapati and Y. Seol and J. Silver and P. R. Desai and S. Das and K. C. Neuman},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.04.749274},
url = {https://www.biorxiv.org/content/10.64898/2026.09.04.749274v1},
note = {CC0},
}
RIS
TY - UNPB
TI - Type II topoisomerase substrate geometry revealed through combined experiment and computation.
AU - M. N. Prajapati
AU - Y. Seol
AU - J. Silver
AU - P. R. Desai
AU - S. Das
AU - K. C. Neuman
PY - 2026
DA - 2026/09/04
PB - bioRxiv
DO - 10.64898/2026.09.04.749274
UR - https://www.biorxiv.org/content/10.64898/2026.09.04.749274v1
AB - Type II topoisomerases (topo IIs) are essential enzymes that regulate DNA topology through a strand-passage mechanism in which a duplex DNA (transfer-segment) is passed through a transiently cleaved second duplex DNA (gate-segment). Biochemical and structural approaches have revealed critical details of the binding and cleavage of the gate-segment DNA. However, capture of the transfer-segment DNA has proven more difficult to resolve due to the transient nature of the interaction. Nonetheless, selection of the transfer segment with a specific conformation or orientation relative to the gate segment is predicted to govern aspects of topo II activity, including chiral discrimination and the ability to reduce topological complexity below equilibrium. To determine the conformation and orientation of the transfer-segment relative to the gate segment, we combined experimental single-molecule measurements of topo II unlinking a single DNA crossing with Brownian dynamics simulations of the DNA crossing. By correlating the unlinking rate with the geometric features of the DNA crossing, we obtain the complete three-dimensional preferred crossing geometry for strand passage. Strikingly, the preferred crossing geometries for Escherichia coli topoisomerase IV and Methanosarcina mazei topoisomerase VI are distinct and provide structural models of the DNA synapse selected for strand passage along with a mechanistic basis for their differing activities and biological functions. The approach we develop is generalizable, providing unique insights into the kinetic selection of DNA synapse structure.
ER -