The DNA revolution: Crystallography and four-stranded DNA

[Dr. Sarah P. Gurung is a recent PhD graduate from University of Reading who lives in Reading (UK). Sarah originally comes from Balaju, Kathmandu, Nepal and moved to the UK when she was 15 years old.  Her PhD research includes the DNA revolution which is vital towards modern medical treatments. She shares her experience and the main idea of the study with Londonkathmandu.com]

The study and use of DNA has been even more interesting and vital for human treatments and other purposes today. The impossible of yesterday has become possible today. In 1953, Watson and Crick introduced us to their Nobel Prize winning model of DNA. We were shown how the molecule of life comes together as two intertwining strands to form a twisted-ladder structure. This discovery, based on the results obtained by Rosalind Franklin and her PhD student Raymond Gosling, opened doors to numerous fields in science, ranging from medicine to nanotechnology.

The method that the student-supervisor pair used to study DNA is called X-ray Diffraction which is also the technique I used for the majority of my PhD. X-rays, are a form of electromagnetic radiation whose wavelengths are shorter than that of visible light. Therefore it can be used in many day to day applications that are limited to the naked eye, such as in airport security checks and medical CT scans; to see beyond what we cannot see. The German scientist Wilhelm Röntgen is credited for the discovery of X-rays in 1895. He called this then unknown radiation “X-radiation” which has been shortened to X-ray over the years.

Diffraction, on the other hand, is the bending of waves due to encountering an obstacle. It is just like how music coming from a radio in one room will sound different when heard from another room because the sound waves have to go through walls or the water waves will change in direction when you throw a pebble into a pond. Similarly, when X-rays hit an object, which was DNA in the case of Franklin and Gosling, the rays are diffracted or bend to different directions and positions. These positions can then be used as a map to determine the location of each atom within a molecule. This technique can be used to study molecules of a range of sizes, from small to larger biological ones.

The experiments and mathematical calculations conducted by Max von Laue, Paul Peter Ewald and the father-son duo William and Laurence Bragg between1912-1913 gave rise to X-ray Crystallography. They introduced the concept of using crystals as obstacles for X-ray diffraction. Imagine if I fall down a flight of stairs and I need to check if my arm is broken. I would go to a hospital and have an X-ray scan. The report would show me the condition of my solid bones but not that of my blood or other fluids. Similarly, hitting a DNA crystal with X-rays will show how the atoms come together inside. We can also test on DNA in its liquid state using X-rays, but the results will appear more complicated because liquid is free-flowing and it will give us various atomic positions whereas a solid crystal sample will give us more specific positions of the atoms.

For the majority of my project, I worked on how to crystallise or solidify four-stranded DNA for X-ray diffraction studies. A double helix DNA involves pairing of two DNA bases, adenine (A) with thymine (T) and guanine (G) with cytosine (C). This rule is almost natural and prevalent in all living things. However, recent studies have shown that DNA sequences which are rich in G and C can form four-stranded DNA instead of the regular two-stranded structure (Figure 1). Furthermore, these sequences are found in “telomeres” which control the aging process and the promoter regions of cancer genes, suggesting that four-stranded DNA may contribute to regulating cancer. In the past five years as a PhD student and a postdoctoral research assistant, I studied on the factors that affect the stability of these four-stranded DNA. My work heavily focused on C-rich DNA or intercalated “i-motifs” which form more readily in acidic environments. This was initially a topic of great contradiction because the human body has a pH of 7.4 so it was argued that the ‘i-motif’ cannot be present in humans.

However, I managed to crystallise an ‘i-motif’ in pH 7.0, going against the theory that they only form in acidic conditions. Moreover, only just recently, Daniel Christ and his research group in Sydney, Australia, showed the formation of i-motif DNA structures in the nuclei of human cells. Therefore, interest in i-motifs and other four-stranded DNA structures are increasing due to their unique pH-specific properties and their potential regulatory role in the human genome.

In short, my ever-challenging PhD research centred around the idea of solidifying cancer DNA from the human body so that a more precise outcome can be obtained on the DNA’s structure. The results can then be used in the future to check what kinds of drugs/medicines can be used or developed to treat cancer.

 Sarah

Figure 1: (Left) The two-stranded “Double Helix” DNA which is prevalent in most living organisms. The DNA bases A (green) always pairs with T (red) and C (yellow) always pairs with G (blue). (Right) The four-stranded “i-motif” DNA which has the potential to form in cancer genes. It is made up of C DNA bases.

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