title: The C-terminal domain of dnaQ contains the polymerase binding site.
abstract: The Escherichia coli dnaQ gene encodes the 3'-->5' exonucleolytic proofreading (epsilon) subunit of DNA polymerase III (Pol III). Genetic analysis of dnaQ mutants has suggested that epsilon might consist of two domains, an N-terminal domain containing the exonuclease and a C-terminal domain essential for binding the polymerase (alpha) subunit. We have created truncated forms of dnaQ resulting in epsilon subunits that contain either the N-terminal or the C-terminal domain. Using the yeast two-hybrid system, we analyzed the interactions of the single-domain epsilon subunits with the alpha and theta subunits of the Pol III core. The DnaQ991 protein, consisting of the N-terminal 186 amino acids, was defective in binding to the alpha subunit while retaining normal binding to the theta subunit. In contrast, the NDelta186 protein, consisting of the C-terminal 57 amino acids, exhibited normal binding to the alpha subunit but was defective in binding to the theta subunit. A strain carrying the dnaQ991 allele exhibited a strong, recessive mutator phenotype, as expected from a defective alpha binding mutant. The data are consistent with the existence of two functional domains in epsilon, with the C-terminal domain responsible for polymerase binding.
authors.affiliation: Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709, USA.
authors.forename: S A
authors.initials: SA
authors.lastname: Taft-Benz
authors.forename: R M
authors.initials: RM
authors.lastname: Schaaper
mesh_terms: D001665:Binding Sites; D004258:DNA Polymerase III / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D043211:Exodeoxyribonuclease V; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics
chemical_list: D029968:Escherichia coli Proteins; D010446:Peptide Fragments; D004258:DNA Polymerase III; C468316:dnaQ protein, E coli; D005090:Exodeoxyribonucleases; D043211:Exodeoxyribonuclease V