Short deoxyribonucleic acid repair patch length in Escherichia coli is determined by the processive mechanism of deoxyribonucleic acid polymerase I.

Abstract:

The lengths of ultraviolet irradiation-induced repair resynthesis patches were measured in repair-competent extracts of Escherichia coli. Extracts containing wild-type deoxyribonucleic acid (DNA) polymerase I introduced a patch 15 to 20 nucleotides in length during repair of ColE1 plasmid DNA; extracts containing the polA5 mutant form of DNA polymerase I introduced a patch only about 5 nucleotides in length in a similar reaction. The repair patch length in the presence of either DNA polymerase corresponded to the processivity of that polymerase (the average number of nucleotides added per enzyme-DNA binding event) as determined with purified enzymes and DNA treated with a nonspecific endonuclease. The base composition of the repair patch inserted by the wild-type DNA polymerase was similar to that of the bacterial genome, whereas the patch inserted by the mutant enzyme was skewed toward greater pyrimidine incorporation. This skewing is expected, considering the predominance of pyrimidine incorporation occurring at the ultraviolet lesion and the short patch made by the mutant enzyme. Since the defect in the polA5 DNA polymerase which causes premature dissociation from DNA is reflected exactly in the repair patch length, the processive mechanism of the polymerase must be a central determinant of patch length.

Polymerases:

Topics:

Source / Purification, Exonuclease Activity, Nucleotide Incorporation, Kinetic Parameters

Status:

new topics/pols set partial results complete validated

Results:

Polymerase Reference Property Result Context
Eco Pol I Matson SW1981 Cloned or native Native organism
Eco Pol I Matson SW1981 5-3' Exonuclease Yes
Eco Pol I Matson SW1981 Tagged No
Eco Pol I Matson SW1981 Full length or truncated Full length
Eco Pol I Matson SW1981 Processivity 20bp
Eco Pol I Matson SW1981 KM 2uM Reaction: Nucleotide incorporation; Substrate: dNTPs
Eco Pol I Matson SW1981 Nick Extension Yes, via strand displacement

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