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The repair patch of E. coli (A)BC excinuclease.
Sibghat-Ullah1, A Sancar, J E Hearst
1Department of Biochemistry and Biophysics, University of North Carolina School of Medicine, Chapel Hill 27599.
Nucleic Acids Research
|September 11, 1990
Summary
DNA repair patch size was determined using E. coli DNA polymerase I (Poll) and (A)BC excinuclease. The study found the repair patch length matched the 12-nucleotide gap created by the excision nuclease.
Area of Science:
- Molecular Biology
- DNA Repair Mechanisms
- Enzymology
Background:
- Thymine-psoralen monoadducts are DNA lesions that require excision and repair.
- E. coli DNA polymerase I (Poll) and (A)BC excinuclease are key enzymes in DNA repair pathways.
- Understanding DNA repair patch size is crucial for assessing the fidelity of these processes.
Purpose of the Study:
- To determine the precise size of the DNA repair patch synthesized by E. coli DNA polymerase I (Poll).
- To investigate the length of the repair patch following the removal of thymine-psoralen monoadducts by (A)BC excinuclease.
Main Methods:
- Utilized M13mp19 DNA containing a single psoralen monoadduct.
- Employed (A)BC excinuclease for excision, followed by Poll for repair synthesis using dNTPs with phosphorothioate linkages.
- Analyzed repair patch size by preferential hydrolysis of phosphorothioate bonds.
Main Results:
- The repair process resulted in a patch containing phosphorothioate linkages.
- Hydrolysis of these linkages revealed a repair patch size equivalent to the 12-nucleotide gap created by the excision nuclease.
- Minimal nick translation was observed during the repair synthesis.
Conclusions:
- The repair patch synthesized by E. coli Poll precisely fills the gap left by (A)BC excinuclease.
- This study quantifies the nucleotide incorporation during base excision repair, confirming enzyme processivity.
- The findings contribute to understanding the accuracy and efficiency of DNA repair in E. coli.