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Title: Biochemical and structural characterisation of human telomeric complexes
Author: Court, R. I.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2004
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In humans there are two proteins that specifically bind the duplex telomeric DNA, human TTAGGG Repeat Binding Factor 1 and 2 (hTRF1 and hTRF2). Although similar in sequence and architecture, hTRF1 and hTRF2 play different roles. hTRF1 regulates telomere length while hTRF2 acts to cap telomere ends and prevents them being recognised as DNA breaks. This work has used biochemical and structural techniques to gain insights into three telomeric complexes, hTRF2 and its cofactor hRap1 and hTRF1 and hTRF2 DNA-binding domains in complex with telomeric DNA. The first part of this thesis covers the characterisation of the interaction between hTRF2 and its partner hRap1. Minimal interaction domains of hTRF2 and hRap1 were successfully defined. Constructs were then designed for use in crystallisation trials, but as yet no crystals have been attained. The second part of this thesis involves the interactions of hTRF1 and hTRF2 with telomeric DNA. Biochemical studies were carried out with the DNA-binding domains to compare binding activity. Then, hTRF1 and hTRF2 DNA-binding domains were co-crystallised with telomeric DNA fragments and their structures determined at high resolution. The structures of the two complexes are highly similar, but there are subtle differences in the details of DNA binding. The structures show that much of the sequence specificity of hTRF1 and hTRF2 is conferred by the presence of water molecules at the protein-DNA interface and that specific sequence recognition as well as binding of duplex telomeric DNA is conserved from yeasts to mammals.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available