Genetics

• Experiments that concluded DNA is a genetic material

  
• 1928 - Fred Griffith - Streptococcus pneumoniae


  1. Used two strains: virulent (S strain) and non-virulent (R strain)

  2. Injected mice with a mixture of living R strain and heat-killed S strain

  3. Mice developed pneumonia and died

  4. Hypothesis - bacteria could do 'transformation'; suggested that DNA could transfer traits between different strains
• 1944 - Oswald Avery, Colin MacLeod, Marlyn McCarty


  1. Separated cell components like DNA, RNA and proteins from heat-killed bacteria

  2. Non-virulent R strain was mixed with each of the components to observe transformation

  3. Observation - when DNA was degraded using deoxyribonuclease, transformation did not occur, when RNA was degraded (ribonuclease), transformation did occur

  4. Conclusion - DNA is the transforming principle

  
• 1952 - Alfred Hershey, Martha Chase


  1. Used T2 bacteriophage which has a protein coat surrounding DNA

  2. Protein coat labelled with radioactive sulphur - 35S

  3. DNA labelled with radioactive phosphorus - 32P

  4. Result - only 32P found in infected bacteria and in phage progeny, 35S only found in phage 'ghosts' that failed to enter the bacteria

  5. This confirmed DNA as genetic material
• Ribose


  Sugar found in RNA
• Deoxyribose


  Sugar found in DNA, does not have an oxygen atom at 2' carbon

  
• Nucleic acids


  Types - DNA and RNA (polynucleotides)

  
• Nucleic acids store and transmit genetic information
• Nitrogenous bases in DNA

  • Adenine and guanine (purines) have two nitrogen rings
  • Cytosine, thymine, and uracil (pyrimidines) have one nitrogen ring
  • Bases are attached to sugar via glycosidic bond
  • Planar rings, usually uncharged under physiological conditions

  
• Tautomeric form of nitrogenous bases is rare (<0.01%)
• Tautomers can result in DNA replication errors and genetic variation

  
• Human genome = 3Gbp = 3 x10^9 base pairs

  
• Approximately 100,000 bp = 0.0033% are rare tautomers
• Nucleoside


  Base + sugar
• Nucleotide


  Nucleoside + phosphate
• Functions of nucleotides


  • ATP acts as energy storage and phosphoryl donor
  • CoA - acetyl group activation and transfer, can also act as donor
  • S-adenosyl methionine - methyl group donor
  • NAD+/NADH or NADP+/NADPH - involved in oxidation-reduction reactions
  • FAD/FADH2 - oxidation-reduction mechanisms
• Linkage of nucleotides in a polynucleotide chain

  1. Monomer - nucleotide

  2. Written from 5' to 3'

  3. Joined by phosphodiester bond between the hydroxyl group at 3' end of one sugar and the phosphate attached to 5' hydroxyl of next sugar

  4. Repeating unit of nucleotide chain - sugar phosphate backbone

  
• Contributions for structure of DNA

  • Chargaff - composition of DNA
  • Rosalind Franklin and Maurice Wilkins - structure of DNA
  • Watson and Crick - proposed double helix model

  
• Chargaff's rules


  [G] = [C] and [A] = [T] (number of pyrimidines = number of purines)
• Organisms with higher temperature have more % G+C
• Rosalind Franklin and Maurice Wilkins' findings

  • Used X-ray diffraction analysis of DNA fibres
  • Photo 51 showed X-shaped diffraction pattern suggested helical structure composed of two intertwined helices
  • 34 Å (3.4 nm) and approx. 10 bp/turn
  • 3.4 Å (0.34 nm) rise per bp
  • Helix diameter 20 Å (2 nm)

  
• 1953 - Watson and Crick proposed double helix model of DNA with complementary base pairing
• Watson and Crick's three-dimensional model of DNA

  • Two antiparallel strands twisted around each other to form double helix
  • Strands run in opposite directions, one in 5' to 3' and other in 3' to 5' direction
  • Helix has a right-handed twist (clockwise) with two strands spiralling around a central axis
  • Nitrogenous bases are paired in the interior of helix, A pairs with T and G pairs with C
  • Hydrophilic sugar-phosphate backbone forms outer structural framework
  • Major groove - wide and deep, minor groove - narrow and shallow

  
• Chemical bonds maintaining DNA structure

  • H-bonds between complementary base pairs (2 H-bonds A-T, 3 H-bonds G-C)
  • Phosphodiester bonds linking sugar molecules in adjacent nucleotides
  • Van der Waals between adjacent base pairs

  
• Comparison of A, B and Z-DNA

  • B-DNA - predominant configuration in cells, helix diameter 2nm, 10.5 bp per turn
  • A-DNA - more squashed appearance, diameter 2.6nm, 11 bp per turn
  • Z-DNA - alternating pyrimidines and purines, left-handed helix, diameter 1.8nm, 12 bp per turn

  
• Importance of DNA coiling

  • DNA has topological variations such as supercoiling
  • Supercoiling allows for DNA packaging and fitting into limited space
  • Influences accessibility of DNA to transcription factors, RNA polymerase, and other regulatory proteins
  • Affects stability and accessibility during replication and repair

  
• Generating a supercoil


  1. Cut a circular molecule and hold one end while twisting the other end

  2. Reattach the ends - DNA twists to restore preferred number of bases per turn

  3. Causes DNA to wrap around itself as coil

  
• Negative supercoiling


  Twists in helix are fewer than twists in relaxed state, most common form

  
• Positive supercoiling


  Overwound DNA double helix, more twists in coiled form than relaxed