Controlled rotation mechanism of DNA strand exchange by the Hin serine recombinase

Botao Xiao; Meghan M. McLean; Xianbin Lei; John F. Marko; Reid C. Johnson

doi:10.1038/srep23697

Controlled rotation mechanism of DNA strand exchange by the Hin serine recombinase

Botao Xiao, Meghan M. McLean, Xianbin Lei, John F. Marko, Reid C. Johnson^*

^*Corresponding author for this work

Molecular Biosciences

Research output: Contribution to journal › Article › peer-review

6 Scopus citations

Abstract

DNA strand exchange by serine recombinases has been proposed to occur by a large-scale rotation of halves of the recombinase tetramer. Here we provide the first direct physical evidence for the subunit rotation mechanism for the Hin serine invertase. Single-DNA looping assays using an activated mutant (Hin-H107Y) reveal specific synapses between two hix sites. Two-DNA â œ braidingâ experiments, where separate DNA molecules carrying a single hix are interwound, show that Hin-H107Y cleaves both hix sites and mediates multi-step rotational relaxation of the interwinding. The variable numbers of rotations in the DNA braid experiments are in accord with data from bulk experiments that follow DNA topological changes accompanying recombination by the hyperactive enzyme. The relatively slow Hin rotation rates, combined with pauses, indicate considerable rotary friction between synapsed subunit pairs. A rotational pausing mechanism intrinsic to serine recombinases is likely to be crucial for DNA ligation and for preventing deleterious DNA rearrangements.

Original language	English (US)
Article number	23697
Journal	Scientific reports
Volume	6
DOIs	https://doi.org/10.1038/srep23697
State	Published - Apr 1 2016

Funding

Research at NU was supported by the NSF through grants MCB-1022117 and DMR-1206868, and by the NIH through grants R01-GM105847 and U54-CA193419. Research at UCLA was supported by NIH grant R01-GM38509. Work at HUST was supported by the National Science Foundation of China (11372116) and the Fundamental Research Funds for the Central Universities (HUST 0118012051). We thank Dr. Hua Bai and Dr. John Graham for technical assistance and helpful discussions.

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title = "Controlled rotation mechanism of DNA strand exchange by the Hin serine recombinase",

abstract = "DNA strand exchange by serine recombinases has been proposed to occur by a large-scale rotation of halves of the recombinase tetramer. Here we provide the first direct physical evidence for the subunit rotation mechanism for the Hin serine invertase. Single-DNA looping assays using an activated mutant (Hin-H107Y) reveal specific synapses between two hix sites. Two-DNA {\^a} {\oe} braiding{\^a} experiments, where separate DNA molecules carrying a single hix are interwound, show that Hin-H107Y cleaves both hix sites and mediates multi-step rotational relaxation of the interwinding. The variable numbers of rotations in the DNA braid experiments are in accord with data from bulk experiments that follow DNA topological changes accompanying recombination by the hyperactive enzyme. The relatively slow Hin rotation rates, combined with pauses, indicate considerable rotary friction between synapsed subunit pairs. A rotational pausing mechanism intrinsic to serine recombinases is likely to be crucial for DNA ligation and for preventing deleterious DNA rearrangements.",

author = "Botao Xiao and McLean, {Meghan M.} and Xianbin Lei and Marko, {John F.} and Johnson, {Reid C.}",

note = "Funding Information: Research at NU was supported by the NSF through grants MCB-1022117 and DMR-1206868, and by the NIH through grants R01-GM105847 and U54-CA193419. Research at UCLA was supported by NIH grant R01-GM38509. Work at HUST was supported by the National Science Foundation of China (11372116) and the Fundamental Research Funds for the Central Universities (HUST 0118012051). We thank Dr. Hua Bai and Dr. John Graham for technical assistance and helpful discussions.",

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AU - Xiao, Botao

AU - McLean, Meghan M.

AU - Lei, Xianbin

AU - Marko, John F.

AU - Johnson, Reid C.

N1 - Funding Information: Research at NU was supported by the NSF through grants MCB-1022117 and DMR-1206868, and by the NIH through grants R01-GM105847 and U54-CA193419. Research at UCLA was supported by NIH grant R01-GM38509. Work at HUST was supported by the National Science Foundation of China (11372116) and the Fundamental Research Funds for the Central Universities (HUST 0118012051). We thank Dr. Hua Bai and Dr. John Graham for technical assistance and helpful discussions.

PY - 2016/4/1

Y1 - 2016/4/1

N2 - DNA strand exchange by serine recombinases has been proposed to occur by a large-scale rotation of halves of the recombinase tetramer. Here we provide the first direct physical evidence for the subunit rotation mechanism for the Hin serine invertase. Single-DNA looping assays using an activated mutant (Hin-H107Y) reveal specific synapses between two hix sites. Two-DNA â œ braidingâ experiments, where separate DNA molecules carrying a single hix are interwound, show that Hin-H107Y cleaves both hix sites and mediates multi-step rotational relaxation of the interwinding. The variable numbers of rotations in the DNA braid experiments are in accord with data from bulk experiments that follow DNA topological changes accompanying recombination by the hyperactive enzyme. The relatively slow Hin rotation rates, combined with pauses, indicate considerable rotary friction between synapsed subunit pairs. A rotational pausing mechanism intrinsic to serine recombinases is likely to be crucial for DNA ligation and for preventing deleterious DNA rearrangements.

AB - DNA strand exchange by serine recombinases has been proposed to occur by a large-scale rotation of halves of the recombinase tetramer. Here we provide the first direct physical evidence for the subunit rotation mechanism for the Hin serine invertase. Single-DNA looping assays using an activated mutant (Hin-H107Y) reveal specific synapses between two hix sites. Two-DNA â œ braidingâ experiments, where separate DNA molecules carrying a single hix are interwound, show that Hin-H107Y cleaves both hix sites and mediates multi-step rotational relaxation of the interwinding. The variable numbers of rotations in the DNA braid experiments are in accord with data from bulk experiments that follow DNA topological changes accompanying recombination by the hyperactive enzyme. The relatively slow Hin rotation rates, combined with pauses, indicate considerable rotary friction between synapsed subunit pairs. A rotational pausing mechanism intrinsic to serine recombinases is likely to be crucial for DNA ligation and for preventing deleterious DNA rearrangements.

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Controlled rotation mechanism of DNA strand exchange by the Hin serine recombinase

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