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Nucleic Acids Res. 2016 Dec 1;44(21):10467-10479. Epub 2016 Sep 20.

Dynamic peptides of human TPP1 fulfill diverse functions in telomere maintenance.

Author information

1
Department of Pharmacology, Case Western Reserve University, School of Medicine, Cleveland, OH 44106, USA.
2
Department of Pharmacology, Case Western Reserve University, School of Medicine, Cleveland, OH 44106, USA derek.taylor@case.edu.
3
Department of Biochemistry, Case Western Reserve University, School of Medicine, Cleveland, OH 44106, USA.

Abstract

Telomeres are specialized nucleoprotein complexes that comprise the ends of linear chromosomes. Human telomeres end in a short, single-stranded DNA (ssDNA) overhang that is recognized and bound by two telomere proteins, POT1 and TPP1. Whereas POT1 binds directly to telomere ssDNA, its interaction with TPP1 is essential for localization of POT1 to the telomere. TPP1 also provides enhanced binding and sequence discrimination that regulates POT1-TPP1 interactions exclusively with telomere ssDNA. Finally, TPP1 recruits telomerase, the enzyme responsible for synthesis of telomere DNA, to the telomere. While the oligosaccharide-oligonucleotide-binding (OB)-fold domain of TPP1 has been solved by X-ray crystallography, the molecular interactions within the POT1-TPP1-ssDNA ternary complex and the conformational changes that contribute to its diverse functions remain ambiguous. We employed hydrogen/deuterium exchange combined with mass spectrometry to identify three peptides, all residing within the OB-fold of TPP1, that exhibit altered exchange rates upon complex formation or ssDNA binding. Mutation of these regions combined with functional assays revealed the diverse contributions of each moiety in protein-protein interactions, regulating telomerase activity or DNA-binding. Together, these functional data combined with biophysical analyses and homology modeling provide a molecular understanding of the diverse contributions of TPP1 in telomere maintenance.

PMID:
27655633
PMCID:
PMC5137443
DOI:
10.1093/nar/gkw846
[Indexed for MEDLINE]
Free PMC Article

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