title: Domain mapping of Escherichia coli RecQ defines the roles of conserved N- and C-terminal regions in the RecQ family.
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.
authors.affiliation: Department of Biomolecular Chemistry, 550 Medical Science Center, 1300 University Avenue, University of Wisconsin, Madison, WI 53706-1532, USA.
authors.forename: Douglas A
authors.initials: DA
authors.lastname: Bernstein
authors.forename: James L
authors.initials: JL
authors.lastname: Keck
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D017124:Conserved Sequence; D004247:DNA / Q000378:metabolism; D004265:DNA Helicases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D010450:Endopeptidases / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology*; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D053484:RecQ Helicases; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017384:Sequence Deletion
chemical_list: D011994:Recombinant Proteins; D004247:DNA; D010450:Endopeptidases; D000251:Adenosine Triphosphatases; C506725:RECQL protein, human; C506724:RecQ protein, E coli; D004265:DNA Helicases; D053484:RecQ Helicases