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A single residue unique to DinB‐like proteins limits formation of the Pol IV multi‐protein complex in Escherichia coli
Author(s) -
Cafarelli Tiziana Moccia,
Rands Thomas J.,
Benson Ryan W.,
Rudnicki Pamela A.,
Lin Ida,
Godoy Veronica G.
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.538.7
Subject(s) - genetics , biology
The mechanisms which regulate the activity of DinB, the most evolutionary conserved error‐prone Y‐family DNA polymerase, are only beginning to be understood. We focused on the regulation of DinB by protein‐protein interactions. We identified two conserved residues, Cysteine 66 (C66) and Proline 67 (P67), which are predicted to be non‐catalytic and are unique among DinB‐like proteins. We constructed DinB(C66A) and found that it co‐purifies with its interacting partners, RecA and UmuD, to a greater extent than DinB. We confirmed in vitro that DinB(C66A) has a higher affinity for RecA and UmuD relative to DinB, but note that the increased affinity of DinB(C66A) for UmuD is RecA‐dependent. Expression of dinB(C66A) from the chromosome resulted in detectable differences in lesion bypass fidelity and homologous recombination. The study of DinB(C66A) has revealed a key interface, which modulates the strength of interaction, and has suggested a binding order of RecA and UmuD to DinB. Uncovering the regulatory mechanisms of Y‐family DNA polymerases will permit their manipulation to deter acquisition of antibiotic resistance and to gain insights into cancer development.