Schizosaccharomyces pombe Protection of Telomeres 1 Utilizes Alternate Binding Modes To Accommodate Different Telomeric Sequences

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• 作者：Sarah E. Altschuler ; Thayne H. Dickey ; Deborah S. Wuttke
• 刊名：Biochemistry
• 出版年：2011
• 出版时间：September 6, 2011
• 年：2011
• 卷：50
• 期：35
• 页码：7503-7513
• 全文大小：407K
• 年卷期：v.50,no.35(September 6, 2011)
• ISSN：1520-4995

文摘

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鈥?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)₂GGT, 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鈥?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.