ATPase/helicase motif mutants of Escherichia coli PriA protein essential for recombination‐dependent DNA replication

Backgrounds: PriA protein, a DEXH‐type helicase with C2C2 zinc‐finger motifs, plays essential roles in RecA‐dependent modes of Escherichia coli chromosomal DNA replication, namely inducible and constitutive stable DNA replication (iSDR and cSDR respectively, which may be initiated from a D‐loop or R...

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Bibliographic Details
Published in:Genes to cells : devoted to molecular & cellular mechanisms 2003-03, Vol.8 (3), p.251-261
Main Authors: Tanaka, Taku, Taniyama, Chika, Arai, Ken‐Ichi, Masai, Hisao
Format: Article
Language:English
Subjects:
Adenosine Triphosphatases - genetics
Adenosine Triphosphatases - metabolism
Amino Acid Motifs - genetics
Amino Acid Motifs - physiology
DNA - metabolism
DNA Helicases - genetics
DNA Helicases - metabolism
DNA Replication - genetics
DNA Replication - physiology
DNA-Binding Proteins - genetics
DNA-Binding Proteins - metabolism
Escherichia coli - enzymology
Escherichia coli - genetics
Mutation
Replication Protein A
Structure-Activity Relationship
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Summary:Backgrounds: PriA protein, a DEXH‐type helicase with C2C2 zinc‐finger motifs, plays essential roles in RecA‐dependent modes of Escherichia coli chromosomal DNA replication, namely inducible and constitutive stable DNA replication (iSDR and cSDR respectively, which may be initiated from a D‐loop or R‐loop structure), and in repair of double‐stranded DNA breaks generated by various genotoxic agents or spontaneously during the course of DNA replication. However, the roles of ATPase/DNA helicase activities in functions of PriA are not well understood. Results: We have generated and characterized mutants of PriA protein carrying amino acid substitutions in its conserved ATPase/DNA helicase motifs, namely the Walker A, B and QXXGRXGR motifs. All these mutants were deficient in ATP hydrolysis and DNA helicase activities, but showed wild‐type levels of D‐loop DNA binding, except for the Walker B mutant which showed reduced DNA binding activity, suggesting that the helicase motifs are not directly involved in the DNA binding activity of PriA protein. They also rescued the low viability and UV‐sensitivity of priA null cells. However, they did not rescue iSDR or cSDR—alternative modes of chromosomal DNA replication of the E. coli genome dependent on recombination functions—to the full extent. Conclusions: ATPase/DNA helicase activities of PriA protein are required for full‐level DNA synthesis in recombination‐dependent modes of DNA replication in E. coli.
ISSN:1356-9597
1365-2443
DOI:10.1046/j.1365-2443.2003.00630.x