Introduction, Evolution and Genomes-7 Flashcards
DNA structure
What is the chemical nature of DNA?
1930.
Showed that each building block of DNA is a nucleotide (a phosphate group linked to a deoxyribose sugar- which is linked to 4 nitrogenous bases).
What did Pheobus Levene propose?
1909.
He has presented his ‘tetranucleotide theory’ before he had even worked out the structure of a nucleotide.
Describe the sugar in DNA?
Pentose (5 carbon) deoxyribose.
Similar to ribose but lacks the -OH on 2-prime.
By convention, the C atoms are numbered 1’ to 5’.
Describe the nitrogenous bases in DNA?
Adenine and Guanine look like purine (5C and 6C ring attached). Adenine has an amine group on C6 whereas Guanine has a double bond O.
Thymine, cytosine and uracil are pyrimidines. Thymine has a methyl group on C1 whereas Uracil has a H. Cytosine is the same as Uracil except it has an amine group on C6.
How are nucleosides named?
the C number of the C-N bond, deoxy and the name of the base.
What are the bonds in nucleotides?
Glycosidic bonds- between sugar C1’ and N9 (purine) or N1 (pyrimidine).
Ester bond- between sugar 5C’ and the phosphate.
What is the nomenclature of DNA?
Base, nucleoside, nucleotide (monophosphate, diphosphate and triphosphate).
How is DNA read and why?
DNA is written and read 5’ to 3’.
DNA is a polynucleotide with polarity (directionality), the 5’ end has a phosphate group and the 3’ end has a hydroxyl group.
A phosphodiester bond links the 3’ C of one nucleotide to the 5’ C of the next.
You must consider the directionality of a nucleic acid due to the importance of DNA polarity.
What was the model Pheobus Levene presented?
Tetranucleotide model.
1910- worked on the model.
1930- presented the model.
Four nucleotides occurred in tetranucleotide blocks with bases pointing outwards.
Why was the DNA transforming principle not believed?
The tetranucleotide model suggests that DNA was simple and repetitive and could not be the genetic material.
Avery et al were not believed (1944). Hershey and Chase could not go further to state that DNA has some function (1952).
If Levene’s tetranucleotide model was correct then what would the proportion of nucleotides be in DNA?
%T=%A=%G=%C
What were Chargaff’s results for the proportion of nucleotides present in DNA?
In 1950 he measured the base composition in different organisms.
%T=%A and %G=%C (%GC varies with organism)
What clues led to Linus Pauling’s Nobel Prize?
In 1951, he described the alpha helix, a structural motif present in many proteins.
He used X-ray crystallography.
The cross formed by the ‘reflections’ gave him the clue.
What is X-ray crystallography?
The crystalline target molecule diffracts X-rays and cause exposed patches (reflections) on photographic films.
The resulting diffraction pattern is a unique signature of the molecule.
What is the wave theory of light?
1678.
When light passes through a small opening, comparable in size to the wavelength of light, a wave front is propagated on the other side. A single spot appears on a screen.
Huygens showed that if the slit is wider, all points across the slid act as a point source. The result is a single slit diffraction pattern on the screen.
If there are two slits, the diffraction patterns interfere. The result is a complex double slit interference pattern on the screen.
What is diffraction effected by?
Width, how many slits and how far they are.
How does the diffraction grating effect the image?
Produces a sharper image and reduces the intermediate peaks to invisibility.
One can calculate how far apart the slits are if one knows the wavelength of the incident light.
How does diffraction around solid objects look like?
1818- the spot, concentrated light in the middle of a shadow.
Augustin Fresnel showed, diffraction also occurred around a solid object with the same width as a slit.
If there are two solid objects, the diffraction patterns interfere.
What images do gratings and grids produce?
Vertical gratings- horizontal spots.
Horizontal gratings- vertical spots.
Grids- give a cross.
Angled grids- an angled cross.
How does spatial separation of points affect an image?
Alters the interference fringes.
Features that are close produce widely separated reflections and features that are distant produce closely separated reflections.
Why do reflections from a helix form a cross?
Aligned helices effectively form a diffraction grid.
What theory explains why reflections from a helix form a cross?
The wave theory of light.
How were images of DNA produced?
DNA could not be crystallised at the time, but it could be stretched into long fibres that could be mounted in front of the X-ray source (X-ray fibre diffraction).
Maurice Wilkins stretched DNA and air dried it.
Raymond Gosling, Rosalind Franklin’s PhD student stretched DNA and left it hydrated.
What clues led to the Nobel Prize for X-ray images of DNA?
1950- Maurice Wilkins produced an X-ray diffraction of dried DNA (in the A form). It was high resolution, but without a clear model in mind, it was difficult to interpret.
1952- after 100hrs of exposure, Gosling and Franklin produced ‘Photo 51’ using hydrated DNA (in the B form). The cross is unmistakable- DNA must be helical.
Who were the key players in the race for finding the structure of DNA?
Linus Pauling- proposed a triple helix 3-strand model, with the bases pointing outwards.
Rosalind Franklin- generated X-ray diffraction patterns of hydrated DNA.
James D Watson, Francis H Crick and Maurice HF Wilkins- proposed a double helix 2-strand model, with the bases pointing inwards.
What is photo 51?
Allowed the dimension of B-DNA to be deduced:
The height of one helical turn- each turn of the helix was measured to be 3.4nm. The closer the spots, the larger the actual distance in the target- therefore the close horizontal bars correspond to large features (the helical turns).
The separation of the bases- there were thought to be 10 bases per turn of the helix. The more distant the spots, the smaller the actual distance in the target. The distance from the middle of the pattern to the edge gives the stacking of the bases (~0.34nm).
The pitch of the helix- the data consisted with a helix with a 2 nm diameter.
What did Linus Pauling publish in 1953?
A triple-helix model of DNA.
He did not have access to Franklin’s photo 51.
The sugar-phosphate backbones faced inwards and the nitrogenous bases faced out.
What clues led to a Nobel Prize of DNA structure?
A deduction from photo 51- James Watson had access to Rosalind Franklin’s photo 51. The data was consistent with a double helix, with paired bases.
Chargaff’s rules- discovering the DNA structure. 2 purines = too wide, 2 pyrimidines = too narrow but 1 purine & 1 pyrimidine = perfect (2nm).
Who made a model of DNA and when?
1953.
Watson- specific A/T and G/C base pairing scheme.
Crick- idea of antiparallel strands.
What happened with Erwin Chargaff, molecular biology and the Nobel Prize?
1950- Chargaff’s rules were established. He discussed his results with Crick and Watson.
Early 1950s- Chargaff had doubts about the emerging field of molecular biology.
1962- Crick, Watson and Wilkins received the Nobel Prize. Chargaff withdrew from his lab and complained about his exclusion.
What were the key features of the Watson-Crick model?
Right-handed helix (A and B DNA, not Z).
Bases face in and sugar-phosphate
face out.
Anti-parallel strands. Crick deduced this from a consideration of bond angles, the first confirmation was provided by Arthur Kornberg in 1962 by measuring the ratio of different dinucleotides.
Complementary base-pairing. Bound together by weak hydrogen bonds. A&T (2 H-bonds) and C&G (3 H-bonds).
Base pair distance. Since 1953, refinements have been made. B form has 10.5 bases per turn and helix turn is 3.6 nM.
Major and minor grooves. The sugar phosphate backbones are not equally spaced. Sequence specific DNA binding proteins generally interact with the major groove of B-DNA because the backbones do not get in the way.
What are other structural variants of DNA?
A-DNA: only in low hydration conditions.
B-DNA: the most structurally stable form under physiological conditions.
Z-DNA: taken up physiologically by stretches of altering pyrimidines and purines.
