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Title: The impact of Alu elements on the human proteome
Author: Atkinson, Sarah Elizabeth
Awarding Body: University of Leeds
Current Institution: University of Leeds
Date of Award: 2019
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Approximately 45% of the human genome is comprised of mobile, or transposable, DNA elements (TEs). Of this, 11% is attributed to Alu elements. Alu elements are approximately 300 base pairs in length and are primarily located in the introns of non-coding DNA. However, in some cases, the introduction of an alternative splice site, as a result of an Alu insertion in a protein-coding region, leads to the exonisation of a partial Alu sequence. This exonisation can lead to the expression of an alternative protein isoform which may have disrupted or altered function and therefore, could have the potential to be cause disease. Through the use of bioinformatics, this project firstly aimed to predict the extent of Alu exonisation and subsequent translation in the human proteome. Additionally, through the use of local sequence alignments, the nature of observed insertions could also be studied. Once prior aims were established, a series of techniques were used to study the possible effects of translated Alu insertions on the structure and function of proteins. A number of protein variants were expressed and purified from E. coli. Using biophysical techniques, such as ITC and CD, Alu structure and any effects of Alu insertions on the ligand binding and stability of MBP were studied. Additional binding experiments were performed as a means to explore a potential binding interaction between an Alu-like sequence with geldanamycin, an interaction which was initially observed using phage display. A secondary avenue of research was performed in collaboration with the Aspden and Wurdak groups at the University of Leeds to investigate the difference in translation levels of 'Alu' and 'non-Alu' mRNAs in human cells. Analysis was performed using a combination of polysome profiling, reverse transcription and quantitative PCR.
Supervisor: Taylor, Paul C. ; Webb, Michael E. ; Dilly, Suzanne Sponsor: Valirx plc ; University of Leeds
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available