DNA is Unstable! Luckily your Cells can handle that.

Another Nobel Prize story?! DAMN RIGHT! This time it’s the prize for chemistry, and Tomas Lindahl, Paul Modrich, and Aziz Sancar will collectively bask in the glory for their outstanding work in studying the mechanisms of DNA repair. Given the billions of cell divisions that have occurred in your body between conception and you today, the DNA that is copied each time remains surprisingly similar to the original that was created in the fertilized egg that you once were. Why is that strange? Well from a chemical perspective that should be impossible, with all chemical processes being subject to random errors from time to time. Along with that, DNA is subjected to damaging radiation and highly reactive substances on a daily basis. This should have led to chemical chaos long before you even became a foetus! Now, I would hope that’s not the case for you, so how do our cells prevent this descent into madness? I’ll tell you! It’s because DNA is constantly monitored by various proteins that all work to correct these errors. They don’t prevent the damage from occurring, they just hang around waiting for something to fix, and all three of the winning scientists contributed to our understanding of how our cells achieve this. So! Where do we begin?

A good place to start would be a brief description of the structure of DNA, as this will make things much clearer when we start discussing the research. DNA is primarily a chain of nucleotides, which are themselves made up of three components: a deoxyribose sugar, a phosphate group, and a nitrogenous base. These components are shown bonded together in Figure 1. It is also worth noting that there are four possible bases, each with a slightly different structure, and the one shown in the image is specifically Adenine. The others are known as Thymine, Cytosine, and Guanine, and all attach to the sugar in the same place. The two negative charges on the phosphate group allow it form another bond to the adjacent nucleotide, and this continues on to form a long chain. Two separate chains are then joined together as shown in Figure 2, and voila! A molecule of DNA is formed!

Figure 1: The basic components of DNA. Source: http://pmgbiology.com/2014/10/21/dna-structure-and-function-igcse-a-understanding/
Figure 2: Representation of how the two chains of Nucleotides bond together to form a molecule of DNA. Source: http://www.d.umn.edu/claweb/faculty/troufs/anth1602/pcdna.html
A comparison of Cytosine and its Methylated equivalent. Source: http://blog-biosyn.com/2013/05/15/dna-methylation-and-analysis/

Now that we have a basic understanding of the structure of DNA, the research should make a hell of a lot more sense, and it begins with Tomas Lindahl. In the 1960s, Lindahl found himself asking a question; how stable is our DNA, really? At the time the general consensus among scientists was that it was amazingly resilient. I mean… how else could it remain so constant? If genetic information was in any way unstable, multicellular organisms like us would have never come into existence. Lindahl began his experiments by working with RNA, another molecule found in our cells with a lot of structural similarities to DNA. However, what was surprising was that the RNA rapidly degraded during these experiments. Now it was known that RNA is the least stable of the two molecules, but if was destroyed so easily and quickly, could DNA really be all that stable? Continuing his research, Lindahl demonstrated that DNA does, in fact, have limited chemical stability, and can undergo many reactions within our cells. One such reaction is Methylation, in which a CH3 (methyl) group is added on to one of the bases in the DNA strand. The difference this causes is shown in Figure 3, and can occur with or without the aid of an enzyme. This reaction will become relevant later on, as will the fact that it changes the shape of the base, affecting how other proteins can bind to it. All of these reactions can alter the genetic information stored in DNA, and if they were allowed to persist, mutations would occur much more frequently than they actually do.

Realising that these errors had to be corrected somehow, Lindahl began investigating how DNA was repaired, and by 1986 he had pieced together a molecular image of how “base excision repair” functions. The process involves many enzymes (and I don’t have the time or patience to describe them all), but a certain class known as “DNA glycolsylases” are what actually break the bond between the defective base and the deoxyribose sugar, and the base is removed. Our cells actually contain many enzymes of this type, each of which targets a different type of base modification. Several more enzymes then work together to fill the gap with the correct, undamaged base and there we have it! A mutation has been prevented. To help you visualise all this, you’ll find a graphical representation of it below in Figure 4.

Figure 4: Graphical representation of the process of Base Excision Repair. Source: http://www.nobelprize.org/nobel_prizes/chemistry/laureates/2015/popular-chemistryprize2015.pdf

But the science doesn’t end there folks! Remember, there were three winners, the second of which is Aziz Sancar, who discovered another method of DNA repair. This one is called “nucleotide excision repair”, and involves the removal of entire sets of nucleotides, rather than individual bases. Sancar’s interest was piqued by one phenomenon in particular; when bacteria are exposed to deadly doses of UV radiation, they can suddenly recover if exposed to visible blue light. This was termed “photoreactivation” for… obvious reasons. He was successful in identifying an isolating the genes and enzymes responsible, but it later became clear that bacteria had a second repair mechanism that didn’t require exposure to light of any kind. But Sancar wasn’t about to let these bacteria out-fox him and, after more investigations, he’d managed to identify, isolate, and characterise the enzymes responsible  for this process as well. The bacteria were no match for his chemical prowess!

“But how does it work?!” I hear you shout. Well calm the f**k down and I’ll tell you! UV radiation can be extremely damaging, and can cause two adjacent Thymine bases in a DNA strand to directly bind to each other, which is WRONG! A certain endonuclease enzyme, known as an “exinuclease”, is aware of this wrongness, and decides that this damage must be fixed. It does this by making two incisions on each side of the defect, and a fragment roughly 12 nucleotides long is removed. DNA polymerase and DNA ligase then fill in and seal the gap, respectively, and now we have a healthy strand of bacterial DNA! Sancar later investigated this repair mechanism is humans in  parallel with other research groups, and while it is much more complicated, involving many more enzymes and proteins, it functions very similarly in chemical terms. You want a picture to make it easier? You’ll find it below in Figure 5!

Figure 5: Graphical representation of Nucleotide Excision Repair. Source: http://www.nobelprize.org/nobel_prizes/chemistry/laureates/2015/popular-chemistryprize2015.pdf

The final recipient of the Nobel Prize this year was Paul Modrich, who identified YET ANOTHER repair system (there are loads, you know), which he named the “mismatch repair” mechanism. Early on in his career, Modrich was examining various enzymes that affect DNA, eventually focussing on “Dam Methylase” which couples methyl groups to DNA bases (I TOLD YOU THAT REACTION WOULD BE RELEVANT!). He showed that these methyl groups could basically behave a labels, helping restriction enzymes cut the DNA strand at the right location. But, only a few years earlier, another scientist called Matthew Meselson, suggested that they also indicate which strand to use a template in DNA replication. Working together, these scientists synthesised a virus with DNA that had incorrectly paired bases, and methylated only one of the two DNA strands. When the virus infected, and injected its DNA into the bacteria, the mismatched pairs were corrected by altering the unmethylated strand. It would appear that the repair mechanism recognised the defective strand by the lack of methyl groups. Does it work that way in humans? Probably not. Modrich did manage to map the mismatch repair mechanism in humans, but DNA methylation serves many other functions in human cells, particularly those to do with gene expression and regulation. It is thought that strand-specific “nicks” (lack of a bond between a phosphate group and a deoxyribose sugar) or ribonucleotides (nucleotide components of RNA) present in DNA may direct repair, but the mechanism remains to be found at this point.

Figure 6: Structure of Olaparib. Source: http://www.reagentsdirect.com/index.php/small-molecules/small-molecules-1/olaparib/olaparib-263.html

But why should we care? Granted it is nice to know this stuff (at least I think so), but what can this information be used for? Well, it actually has applications within the world of medicine, as errors in repair mechanisms can often lead to cancer. In many forms of cancer these mechanisms have been at least partially turned off, but the cells are also heavily reliant on the mechanisms that remain active. As we mentioned earlier, a lack of these mechanisms leads to chemical chaos, and that would cause the cancer cells to just die. This has led to drugs designed to inhibit the remaining repair systems to slow down or stop cancer growth entirely! One such drug is Olaparib, and you can see the structure in Figure 6. This drug functions by inhibiting two specific proteins (PARP-1 and PARP-2), which are integral in detecting certain flaws in replicated DNA and directing repair proteins to the site of damage. Cancer cells treated with this drug have been shown to be more sensitive to UV radiation, making one form of treatment much more effective.

And with that, we bring our Nobel Prize stories for this year to an end! I think it’s safe to say that the work described here deserved a prize of some sort, as it not only takes a lot of skill and dedication, but it has led to new medical treatments and a MUCH greater understanding of how our DNA behaves. Have you enjoyed our time spent on the science of the Nobel Prize? DAMN RIGHT YOU HAVE. O_O



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