PMID:9506888
| Citation |
Moolenaar, GF, Franken, KL, van de Putte, P and Goosen, N (1997) Function of the homologous regions of the Escherichia coli DNA excision repair proteins UvrB and UvrC in stabilization of the UvrBC-DNA complex and in 3'-incision. Mutat. Res. 385:195-203 |
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| Abstract |
The nicking of damaged DNA during the nucleotide excision repair reaction in E. coli, is the result of a multi-step process involving three enzymes, UvrA, UvrB and UvrC. The UvrB protein is loaded on the site of the damage by UvrA, forming a stable UvrB-DNA complex. This complex is recognized by UvrC and in the resulting UvrBC-DNA complex dual incision takes place, first on the 3'-side and next on the 5'-side of the damaged nucleotide. A domain in the C-terminal part of UvrB has been identified to be essential for formation of the specific UvrBC-DNA complex that induces the 3'-incision [1]. The N-terminal half of UvrC contains a region that is homologous to this C-terminal domain of UvrB. Using site-directed mutagenesis of a conserved phenylalanine in the homologous regions of UvrB and UvrC two mutants were constructed, UvrB(F652L) and UvrC(F223L). Both proteins were tested in vitro using a DNA substrate with a defined cisplatin lesion. The protein-DNA and protein-protein interactions were studied using bandshift assays and DNAse I footprinting. We show that both domains are important for the binding of UvrC to the UvrB-DNA complex. |
| Links | |
| Keywords |
Adenosine Triphosphate/metabolism; Amino Acid Sequence; Bacterial Proteins/genetics; Bacterial Proteins/metabolism; Bacterial Proteins/physiology; Base Sequence; Cloning, Molecular; DNA Footprinting; DNA Helicases; DNA Repair; DNA, Bacterial/analysis; DNA, Bacterial/genetics; Deoxyribonuclease I/metabolism; Endodeoxyribonucleases; Escherichia coli/genetics; Escherichia coli/metabolism; Escherichia coli Proteins; Molecular Sequence Data; Mutagenesis, Site-Directed; Phenylalanine/genetics; Plasmids; Protein Binding; Sequence Alignment; Sequence Analysis, DNA; Sequence Homology, Amino Acid |
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