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

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Abstract
The τ subunit of Escherichia coli DNA polymerase III holoenzyme interacts with the α subunit through its C-terminal Domain V, τ C 16. We show that the extreme C-terminal region of τ C 16 constitutes the site of interaction with α. The τ C 16 domain, but not a derivative of it with a C-terminal deletion of seven residues (τ C 16Δ7), forms an isolable complex with α. Surface plasmon resonance measurements were used to determine the dissociation constant ( KD ) of the α−τ C 16 complex to be ∼260 pM. Competition with immobilized τ C 16 by τ C 16 derivatives for binding to α gave values of KD of 7 μM for the α−τ C 16Δ7 complex. Low-level expression of the genes encoding τ C 16 and τ C 16▵7, but not τ C 16Δ11, is lethal to E. coli . Suppression of this lethal phenotype enabled selection of mutations in the 3′ end of the τ C 16 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, τ C 24, was found to bind DNA in a salt-sensitive manner while no binding was observed for τ C 16, suggesting that the processivity switch of the replisome functionally involves Domain IV of τ.