title: Schizosaccharomyces pombe protection of telomeres 1 utilizes alternate binding modes to accommodate different telomeric sequences.
abstract: The ends of eukaryotic chromosomes consist of long tracts of repetitive GT-rich DNA with variable sequence homogeneity between and within organisms. Telomeres terminate in a conserved 3'-ssDNA overhang that, regardless of sequence variability, is specifically and tightly bound by proteins of the telomere-end protection family. The high affinity ssDNA-binding activity of S. pombe Pot1 protein (SpPot1) is conferred by a DNA-binding domain consisting of two subdomains, Pot1pN and Pot1pC. Previous work has shown that Pot1pN binds a single repeat of the core telomere sequence (GGTTAC) with exquisite specificity, while Pot1pC binds an extended sequence of nine nucleotides (GGTTACGGT) with modest specificity requirements. We find that full-length SpPot1 binds the composite 15mer, (GGTTAC)(2)GGT, and a shorter two-repeat 12mer, (GGTTAC)(2), with equally high affinity (<3 pM), but with substantially different kinetic and thermodynamic properties. The binding mode of the SpPot1/15mer complex is more stable than that of the 12mer complex, with a 2-fold longer half-life and increased tolerance to nucleotide and amino acid substitutions. Our data suggest that SpPot1 protection of heterogeneous telomeres is mediated through 5'-sequence recognition and the use of alternate binding modes to maintain high affinity interaction with the G-strand, while simultaneously discriminating against the complementary strand.
authors.affiliation: Department of Chemistry and Biochemistry, University of Colorado, Boulder, Colorado 80309-0215, United States.
authors.forename: Sarah E
authors.initials: SE
authors.lastname: Altschuler
authors.forename: Thayne H
authors.initials: TH
authors.lastname: Dickey
authors.forename: Deborah S
authors.initials: DS
authors.lastname: Wuttke
mesh_terms: D001665:Binding Sites / Q000235:genetics; D017124:Conserved Sequence / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D012568:Schizosaccharomyces / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000089804:Shelterin Complex; D034501:Telomere-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D035321:Telomeric Repeat Binding Protein 1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; D029702:Schizosaccharomyces pombe Proteins; D000089804:Shelterin Complex; D034501:Telomere-Binding Proteins; D035321:Telomeric Repeat Binding Protein 1; C499082:pot1 protein, S pombe