Rosalind Franklin and the Structure of DNA

Three men, James Dewey Watson,  Francis Harry Compton Crick and Maurice Hugh Frederick Wilkins, shared the The Nobel Prize in Physiology or Medicine 1962 "for their discoveries concerning the molecular structure of nucleic acids and its significance for information transfer in living material.", that is, they  modeled DNA as a double helix, each strand of the helix has a backbone of  sugar molecules held together by phosphate groups. The two strands are twisted together and held together by hydrogen bonds. But how did they learn what DNA was made up of?

 This is where Rosalind Elsie Franklin enters the story of DNA. In 1951 she was a Research Associate at Kings College London where she worked on  X-ray diffraction studies with her colleague Maurice Wilkins. Her x-ray diffraction images of DNA led to the discovery of the DNA helix. The image on the left is known as "photograph 51" and was an x-ray diffraction image of DNA obtained by Franklin's Ph.D student Raymond Gosling.

X-ray diffraction is an instrumental technique used to elucidate the structure of crystals of chemical compounds. Incoming x-rays are diffracted by the crystal lattice and they exit the crystal at different angles. An x-ray crystallographer like Franklin can measure the angles and intensities of these diffracted x-rays to produce a 3-dimensional picture of the density of electrons in the crystal lattice. The electron density can then be used to determine the locations of atoms within the crystal lattice.

Without Franklin's knowledge, Maurice Wilkins showed this image to James Watson who used it, along with other evidence, to develop a model of DNA. Science historians still debate whether Franklin would have determined the structure of DNA on her own had her images not been shared with Watson.

Rosalind Franklin made important scientific contributions, not only to the discovery of the structure of DNA and RNA, but also in helping us to understand the structure of viruses, coal and graphite.
Unfortunately, Rosalind Franklin died of ovarian cancer in 1958. Nobel Prizes are not generally awarded posthumously so her contribution to the elucidation of the structure of DNA is not well-known.


Further Reading:
Chemistry of DNA
Intramolecular Forces
Intermolecular Forces

Suggested Study Questions:

1. Explain the terms crystalline and amorphous.
2. Give an example of a crystalline substance and an example of an amorphous substance.
3. Explain why DNA had to be crystallised before useful information could be obtained using x-ray diffraction.
4. What does the abbreviation DNA stand for?
5. What are the 4 principle bases that make up DNA?
6. These principle bases occur in pairs; what are these 2 pairs?
7. What kind of chemical bonds act between the atoms making up each base in a strand of DNA?
8. What kind of chemical forces join one of the bases on one strand of DNA to its corresponding pair on the other strand of DNA?
9. If you wanted to separate the 2 strands of a DNA double helix, what sort of chemical bonds would you need to break?
10. If you wanted to separated each base from the backbone of sugar molecules, what sort of chemical bonds would you need to break?

2015 Nobel Prize in (Bio)chemistry?

The 2015 Nobel Prize in Chemistry was awarded to Tomas Lindahl, Paul Modrich and Aziz Sancar for "mechanistic studies of DNA repair".
Cells have developed mechanisms to repair damaged DNA. Four of these mechanisms are:

• photoreactivation


• dark repair (nucleotide excision repair)


• base excision repair


• mismatch repair

Photoreactivation
In the 1920s Hermann Muller found that X-rays could mutate and kill cells.
In the 1940s Albert Kelner found that visible light could stimulate growth recovery after damage caused by UV light, and this was called photoreactivation.
In 1944 Oswald Avery and co-workers showed that DNA is the material of heredity, and in the 1950s it was recognised that DNA became damaged when exposed to UV light.
Renato Dulbecco suggested photoreactivation was an enzymatic reaction dependent on light, which was demonstrated by Stanley Rupert.
In 1978 Aziz Sancar cloned the E. coli photolyase gene, an enzyme responsible for DNA repair in escherichia coli.
In the 1980s Aziz Sancar showed that photolyase can convert the energy of an absorbed photon into chemistry that produces a localised free radical that initiates thymine dimer splitting.

Dark repair (nucleotide excision repair)
In the 1960s Jane Setlow and Richard Setlow showed that thymine dimers inactivated transforming DNA in Hemophilus influenzae and that this was responsible for the biological effect of UV light.
In 1964 Richard Setlow discovered that thymine dimers disappeared from the irradiated, high molecular weight genomic DNA shortly after exposure to UV light and appeared in the low molecular weight fractions, that is, thymine dimers are excised (removed) from the DNA. This mechanism became known as nucleotide excision repair (NER).
In the 1970s Aziz Sancar working with W. Dean Rupp, developed the Maxicell technique for the rapid identification of proteins.
In 1983 Aziz Sancar used purified proteins to reconstitute essential steps in the nucleotide excision repair (NER) pathway, a "cut and patch" method for DNA repair.
Two proteins (UvrA and UvrB) track along the DNA, UvrA recognises damage and causes UvrB to stop tracking and begin unwinding the effected DNA section. Another protein, UvrC, causes the damaged section to be cut out, and then another protein UvrD, causes UvrB to bind and bridge the gap while it is repaired by resynthesising the removed segment via Pol I.

Base excision repair
In the 1970s Tomas Lindahl showed that DNA has limited chemical stability and that modification of the bases of DNA increased the risk of mutations. High levels of spontaneous cytosine deamination leads to the formation of uracil.

Uracil forms base pairs with adenine, so, high levels of cytosine demanination pose a risk of depleting the genetic material from cytosine-guanine base pairs and replacing them with thymine-adenine.
He identified the E. coli uracil-DNA glycosylase (UNG) as the first repair protein which we now know is one member of a large family of proteins that orchestrate base excision repair (BER).
A DNA glycosylase recognises and cuts the base-deoxyribose glycosyl bond of a damaged nucleotide. DNA glycosylase kinks the DNA and the abnormal nucleotide flips out and is removed and the section can then be repaired.

Mismatch repair
During the synthesis of a new DNA strand, a non-Watson-Crick base pair may be formed which distorts the double-stranded DNA helix. These types of errors are known as mismatches.
In 1983 Paul Modrich and Matthew Meselson showed that DNA methylation directed strand-specific elimination of mismatches in E. coli. Modrich developed an assay to isolate the products of the different repair genes and identify the proteins.

Further Reading:
http://www.ausetute.com.au/dna.html
http://www.ausetute.com.au/enzymes.html

Suggested Study Questions:

1. What does the abbrevaition DNA stand for?
2. What do you think when biochemists refer to damaged DNA?
3.  What is meant by the term enzyme?
4. Why do you think enzymes are required in the mechanisms available within a living cell to repair damaged DNA?
5. What is a free radical?
6. Draw the structural formula of thymine.
7. Draw the structural formula for a possible dimer of thymine.
8. What is meant by the term nucleotide?
9. With reference to DNA, what is meant by a base pair?
10. Show how uracil forms a base pair with adenine.
11. Draw the structure of a cytosine-guanine base pair
12. What do you think is meant by the statement, "cytosine deamination leads to the formation of uracil" (structural formulae may be useful in your explanation but you do not need to include chemical reactions).

DNA in Space

Scientists have evidence that some building blocks of DNA, the molecule that carries the genetic instructions for life, found in meteorites were most likely to have been created in space.
The DNA building blocks present in the meteorites are called nucleobases or nucleotide bases, a group of nitrogen-based molecules that are required in the formation of nucleotides, and nucleotides are the molecules that make up DNA. The primary nucleobases in DNA are cytosine, guanine, adenine and thymine.

For the first time, we have three lines of evidence that together give us confidence these DNA building blocks found in meteorites actually were created in space:

1. In two of the meteorites, trace amounts of three molecules related to nucleobases: purine, 2,6-diaminopurine, and 6,8-diaminopurine; were discovered for the first time. 2,6-diaminopurine, and 6,8-diaminopurine are almost never used in biology so these can't be due to contamination from terrestrial sources.
2. The amounts of the two nucleobases adenine and guanine found in DNA, plus hypoxanthine and xanthine which are similar to the nucleobases but not found in DNA, that were found in terrestrial ice samples from near the Antarctic meteorites were much lower, parts per trillion, than in the meteorites, where they were generally present at several parts per billion. This strongly suggests that terrestrial contamination was not responsible for the presence of these molecules in the meteorites.
3. The adenine, guanine, hypoxanthine and xanthine were produced in a completely non-biological reaction. In the lab, these same molecules were generated in non-biological chemical reactions containing hydrogen cyanide, ammonia, and water. This provides a plausible mechanism for their synthesis in the asteroid parent bodies, and supports the notion that they are extraterrestrial

Reference
NASA (2011, August 9). DNA building blocks can be made in space, NASA evidence suggests. ScienceDaily. Retrieved August 11, 2011, from http://www.sciencedaily.com­ /releases/2011/08/110808220659.htm

Further Reading
DNA
Functional Groups

Study Questions

1. What does the abbreviation DNA stand for?
2. Give the accepted abbreviations for each of the following nucleobases:
   • adenine
   • guanine
   • cytosine
   • thymine
3. Draw a structural formula for each of the nucleobases above.
4. On the structural formulae above, circle the functional groups present.
   
5. Below is the structural formula for hypoxanthine:
   
   Which nucleobase is it most similar to? Explain your answer.
6. Guanine can form 3 hydrogen bonds with cytosine. How many hydrogen bonds could be formed between hypoxanthine cytosine. Use a diagram to explain your answer.
7. The structure of 2,6-diaminopurine is shown below:
   
   Circle the two amine functional groups.
8. What functional groups would be found in a molecule of 6,8-diaminopurine?
9. Give the molecular formula for each of the following molecules:
   • hydrogen cyanide
   • ammonia
   • water
10. Why do you think scientists would try to make the nucleobases out of hydrogen cyanide, ammonia, and water in the laboratory?