Telomere resolution by Borrelia burgdorferi ResT through the collaborative efforts of tethered DNA binding domains.
Borrelia burgdorferi, a causative agent of Lyme disease, has a highly unusual segmented genome composed of both circular molecules and linear DNA replicons terminated by covalently closed hairpin ends or telomeres. Replication intermediates of the linear molecules are processed into hairpin telomeres via the activity of ResT, a telomere resolvase. We report here the results of limited proteolysis and mass spectroscopy to identify two main structural domains in ResT, separated by a chymotrypsin cleavage site between residues 163 and 164 of the 449 amino acid protein. The two domains have been overexpressed and purified. DNA electrophoretic mobility shift assays revealed that the C-terminal domain (ResT(164-449)) displays sequence-specific DNA binding to the box 3,4,5 region of the telomere, while the N-terminal domain (ResT(1-163)) exhibits sequence-independent DNA binding activity. Further analysis by DNase I footprinting supports a model for telomere resolution in which the hairpin binding module of the N-terminal domain is delivered to the box 1,2 region of the telomere through its tethering to ResT(164-449). Conversely, ResT(1-164) may play an important regulatory role by modulating both sequence-specific DNA binding activity and catalysis by the C-terminal domain.
Tourand
Yvonne
Y
Department of Biochemistry and Molecular Biology, University of Calgary, Calgary, Alberta, Canada.
Lee
Linda
L
Chaconas
George
G
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D025065:Borrelia burgdorferi / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D002918:Chymotrypsin / Q000378:metabolism; D004261:DNA Replication / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016680:Genome, Bacterial; D008957:Models, Genetic; D011485:Protein Binding; D045522:Recombinases / Q000235:genetics / Q000378:metabolism; D016615:Telomere / Q000201:enzymology / Q000378:metabolism*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D045522:Recombinases; D004706:Endodeoxyribonucleases; D003850:Deoxyribonuclease I; C446515:ResT protein, Borrelia; D002918:Chymotrypsin