Slow unloading leads to DNA-bound ?2-sliding clamp accumulation in live Escherichia coli cells

Moolman, M.C.; Tiruvadi Krishnan, S.; Kerssemakers, J.W.J.; Van den Berg, A.; Tulinski, P.; Depken, M.; Reyes-Lamothe, R.; Sherratt, D.J.; Dekker, N.H.

doi:doi:10.1038/ncomms6820

Slow unloading leads to DNA-bound ?2-sliding clamp accumulation in live Escherichia coli cells

Title

Slow unloading leads to DNA-bound ?2-sliding clamp accumulation in live Escherichia coli cells

Author

Moolman, M.C.
Tiruvadi Krishnan, S.
Kerssemakers, J.W.J.
Van den Berg, A.
Tulinski, P.
Depken, M.
Reyes-Lamothe, R.
Sherratt, D.J.
Dekker, N.H.

Faculty

Applied Sciences

Department

BN/Bionanoscience

Date

2014-12-18

Abstract

The ubiquitous sliding clamp facilitates processivity of the replicative polymerase and acts as a platform to recruit proteins involved in replication, recombination and repair. While the dynamics of the E. coli ?2-sliding clamp have been characterized in vitro, its in vivo stoichiometry and dynamics remain unclear. To probe both ?2-clamp dynamics and stoichiometry in live E. coli cells, we use custom-built microfluidics in combination with single-molecule fluorescence microscopy and photoactivated fluorescence microscopy. We quantify the recruitment, binding and turnover of ?2-sliding clamps on DNA during replication. These quantitative in vivo results demonstrate that numerous ?2-clamps in E. coli remain on the DNA behind the replication fork for a protracted period of time, allowing them to form a docking platform for other enzymes involved in DNA metabolism.

Subject

biological sciences
biophysics
molecular biology

To reference this document use:

http://resolver.tudelft.nl/uuid:8e45480a-a59b-4ac9-88f7-8a09118e875b

DOI

https://doi.org/10.1038/ncomms6820

Publisher

Nature Publishing Group

ISSN

2041-1723

Source

Nature Communications, 5, 2014