The unstructed C-terminus of the τ subunit of Escherichia coli DNA polymerase III holoenzyme is the site of interaction with the α subunit

Slobodan Jergic, Kiyoshi Ozawa, Neal K. Williams, Xun Cheng Su, Daniel D. Scott, Samir M. Hamdan, Jeffrey A. Crowther, Gottfried Otting, Nicholas E. Dixon*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

44 Scopus citations

Abstract

The τ subunit of Escherichia coli DNA polymerase III holoenzyme interacts with the α subunit through its C-terminal Domain V, τC16. We show that the extreme C-terminal region of τC16 constitutes the site of interaction with α. The τC16 domain, but not a derivative of it with a C-terminal deletion of seven residues (τC16Δ7), forms an isolable complex with α. Surface plasmon resonance measurements were used to determine the dissociation constant (KD) of the α-τC16 complex to be ∼260pM. Competition with immobilized τC16 by τC16 derivatives for binding to α gave values of KD of 7 μM for the α-τC16Δ7 complex. Low-level expression of the genes encoding τC16 and τC16Δ7, but not τC16Δ11, is lethal to E. coli. Suppression of this lethal phenotype enabled selection of mutations in the 3′ end of the τC16 gene, that led to defects in α binding. The data suggest that the unstructured C-terminus of τ becomes folded into a helix-loop-helix in its complex with α. An N-terminally extended construct, τC24, was found to bind DNA in a salt-sensitive manner while no binding was observed for τC16, suggesting that the processivity switch of the replisome functionally involves Domain IV of τ.

Original languageEnglish (US)
Pages (from-to)2813-2824
Number of pages12
JournalNUCLEIC ACIDS RESEARCH
Volume35
Issue number9
DOIs
StatePublished - May 2007
Externally publishedYes

Bibliographical note

Funding Information:
The authors thank Dr Zenta Tsuchihashi for providing plasmid pZT3(1G4G), and Drs Mark Mulcair and Patrick Schaeffer for assistance and advice with SPR experiments. This work was supported by the Australian Research Council, including project grants (to G.O. and N.E.D.), a CSIRO-Linkage Fellowship (to K.O.) and a Federation Fellowship (to G.O.). S.J. held an International Postgraduate Research Award. Funding to pay the Open Access publication charge was provided by the University of Wollongong.

ASJC Scopus subject areas

  • Genetics

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