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Xeroderma pigmentosum complementation group A protein acts as a processivity factor
1Department of Pathology, Laboratory Medicine, UMDNJ-New Jersey Medical School, 185 South Orange Avenue, Newark, New Jersey 07103, USA. mlambert@umdnj.edu
Abstract:
We have previously shown that endonucleases present in a protein complex, which has specificity for cyclobutane pyrimidine dimers, locate sites of damage in DNA by a processive mechanism of action in normal human lymphoblastoid cells. In contrast, the endonucleases present in this complex from xeroderma pigmentosum complementation group A (XPA) cells locate damage sites by a distributive or significantly less processive mechanism. Since the XPA protein has been shown to be responsible for the DNA repair defect in XPA cells, this protein was examined for involvement in the mechanism of target site location of these endonucleases. A recombinant XPA protein, produced by expression of the normal XPA cDNA in E. coli, was isolated and purified. The results show that the recombinant XPA protein was able to correct the defect in ability of the XPA endonucleases to act by a processive mechanism of action on UVC irradiated DNA. These studies indicate that the XPA protein, in addition to a role in damage recognition or damage verification, may function as a processivity factor.
Insights
The xeroderma pigmentosum complementation group A (XPA) protein corrects DNA repair defects by enabling endonucleases to locate DNA damage more effectively. This suggests XPA protein acts as a processivity factor in DNA repair.
Area of Science:
- Molecular Biology
- DNA Repair Mechanisms
- Biochemistry
Background:
- Endonucleases in normal cells locate DNA damage via a processive mechanism.
- In xeroderma pigmentosum complementation group A (XPA) cells, this process is distributive or less processive.
- The XPA protein is known to cause the DNA repair defect in XPA cells.
Purpose of the Study:
- To investigate the role of the XPA protein in the target site location mechanism of endonucleases.
- To determine if XPA protein influences the processivity of endonuclease action on damaged DNA.
Main Methods:
- Produced and purified recombinant XPA protein from E. coli.
- Assayed the mechanism of action of XPA endonucleases on UVC-irradiated DNA.
- Compared endonuclease activity in the presence and absence of recombinant XPA protein.
Main Results:
- Recombinant XPA protein restored the processive mechanism of action for XPA endonucleases on damaged DNA.
- The defect in processive DNA damage site location in XPA cells was corrected by the recombinant XPA protein.
Conclusions:
- The XPA protein functions as a processivity factor for endonucleases involved in DNA repair.
- Beyond damage recognition/verification, XPA protein plays a role in ensuring efficient DNA repair site location.