Investigating an Unusual DNA Structure – The I-Motif

Guest Speaker:
Dr John Brazier (School of Pharmacy, University of Reading)
Subject Area:
Pharmacy
Activity:
Seminar
Date & Time:
Wed, 20. January 2016, 14:00 h - 15:00 h
Venue:
N3.04, University of Reading Malaysia - EduCity@Iskandar​

Abstract
The i-motif is a quadruplex structure that is formed by cytosine-rich sequences of DNA and has been found to form in several biologically relevant sequences of DNA. The role of the i-motif in modulating gene expression has recently been elucidated, with the contribution of the loop regions within the structure key to the recognition and binding of transcriptional proteins. The i-motif is of interest for its biological role in gene expression, especially due to its presence in the promoter regions of several gene associated with cancer. It is known from research on biologically derived sequences, that i-motifs formed by different sequences exhibit different thermal and pH stability. It has been reported in the literature that i-motif forming sequences containing longer loops are thermally more stable than those with short loops, with the length of the loop often suggested to be the contributing factor for this stability. Recently we published results that showed unusually high stability for a sequence containing short loops which questioned the need for long loops to impart stability.

To probe this further, we investigated a series of i-motif forming sequences in which we changed the length of the loop regions to between 3 and 8 thymine bases. Thermal UV absorbance experiments, as well as pH titrations showed that i-motifs containing short loops were inherently more stable than those containing long loops. We also showed that both changes to the loops at the bottom of the structure had more effect on the stability than the single loop at the top.

It has been shown that interactions between bases in the loops can stabilise i-motif structures, so we investigated the effect of loop composition on i-motif stability by altering the base sequence of a trimer loop, of the general formula (CCCXYZ)3CCC where X and Z = A,T or G and Y = A, T, C or G. Both UV and pH stability were measured, along with CD spectra at pH 5 and 8. The results showed that base composition lead to differences in stability, but the correlation between thermal and pH stability was approximately linear. The more thermally stable, the more pH stability was present. The most interesting finding was that not all sequences were able to form clearly defined i-motif structures as shown in the CD spectra.

Photo Biography
Sep 2008 – Present: Lecturer in Pharmaceutical Chemistry, University of Reading, UK.
Jul 2006 – Jul 2008: JSPS Post-doctoral Research Fellow, Kyushu University, Japan.
Apr 2003 – Jun 2006: Post-doctoral Researcher, University of Liverpool, UK.
Oct 1999 – Apr 2003: PhD student, University of Sheffield: UK.
Oct 1995 – July 1999: MChem student, University of Warwick, UK.

Dr John Alan Brazier’s research is focussed on understanding DNA secondary structure, and small molecule binding to DNA. This ranges from unusual DNA structures such as the i-motif, to electron transfer processes by metal complexes bound to DNA. He uses a variety of techniques to study these structures and processes, including UV and CD spectroscopy, x-ray crystallography, and transient infra-red spectroscopy.

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