PMID:12771204
| Citation |
Bernstein, DA and Keck, JL (2003) Domain mapping of Escherichia coli RecQ defines the roles of conserved N- and C-terminal regions in the RecQ family. Nucleic Acids Res. 31:2778-85 |
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| Abstract |
RecQ DNA helicases function in DNA replication, recombination and repair. Although the precise cellular roles played by this family of enzymes remain elusive, the importance of RecQ proteins is clear; mutations in any of three human RecQ genes lead to genomic instability and cancer. In this report, proteolysis is used to define a two-domain structure for Escherichia coli RecQ, revealing a large (approximately 59 kDa) N-terminal and a small (approximately 9 kDa) C-terminal domain. A short N-terminal segment (7 or 21 residues) is also shown to be sensitive to proteases. The effects of removing these regions of RecQ are tested in vitro. Removing 21 N-terminal residues from RecQ severely diminishes its DNA-dependent ATPase and helicase activities, but does not affect its ability to bind DNA in electrophoretic mobility shift assays. In contrast, removing the approximately 9 kDa C-terminal domain from RecQ results in a fragment with normal levels of ATPase and helicase activity, but that has lost the ability to stably associate with DNA. These results establish the biochemical roles of an N-terminal sequence motif in RecQ catalytic function and for the C-terminal RecQ domain in stable DNA binding. |
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| Keywords |
Adenosine Triphosphatases/chemistry; Adenosine Triphosphatases/genetics; Adenosine Triphosphatases/metabolism; Amino Acid Sequence; Conserved Sequence; DNA/metabolism; DNA Helicases/chemistry; DNA Helicases/genetics; DNA Helicases/metabolism; Electrophoretic Mobility Shift Assay; Endopeptidases/metabolism; Escherichia coli/enzymology; Molecular Sequence Data; Protein Structure, Tertiary; RecQ Helicases; Recombinant Proteins/metabolism; Sequence Alignment; Sequence Deletion |
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