DNA structure and replication

The Watson and Crick model suggested semi-conservative replication
1. DNA structure suggested replication mechanism
2. knowledge of DNA structure
  1. molecular modelling by Linus Pauling
  2. X-ray diffraction patterns by Rosalind Franklin
    1. DNA helix tightly packed
    2. bases fit together; strands not too far apart
  3. base composition studies by Erwin Chargaff
3. their 1st model - sugar-phosphate strands wrapped around one another with nitrogen bases facing outwards
  1. Rosalind Franklin countered this
  2. nitrogen bases more hydrophobic than sugar-phosphate backbone, so would face inwards
4. tightly packed if:
  1. pyrimidine paired with purine
  2. bases 'upside down' in relation to one another
5. adenine and thymine both structurally and electrically (charges) compatible
6. adenine and guanine paired; 3 hydrogen bonds = structural stability
7. model of complementary base pairing lead to theory
The role of nucleosomes in DNA packing
1. help supercoil DNA
2. Nucelosomes - structures of DNA packaged by histones
  1. central core of 8 histone proteins (octane) with DNA coiled around them
    1. 2 copies of 4 diff types of histones
  2. connected by 'linker' DNA
    1. bound to core by histone molecule, H1
      1. bind to form 30nm fiber; assists packing
  3. facilitates packing of large genomes of eukaryotes
3. association of histones contributes to supercoiling pattern
4. supercoiling allows great length of DNA to be packed in small space in nucleus
The leading strand and the lagging strand
1. leading
  1. continuous replication
  2. continuously following the fork
2. lagging
  1. discontinuous replication
  2. fragments moving away from the replication fork
  3. Okazaki fragements - new fragments created on lagging strand as fork exposed more template strand
3. arranged in anti-parallel fashion
Proteins involved in replication
1. replication carried out by complex enzyme system
  1. helicase unwinds DNA at the fork
  2. topoisomerase releases strain that develops ahead of helicase
2. replication
  1. formation and movement of replication fork
  2. synthesis of leading and lagging strands
3. single stranded binding proteins keep strands apart long enough for template strand to be copied
4. RNA primer needed to start replication
  1. created by DNA primase
  2. necessary to initiate DNA polymerase activity
5. many primers on lagging strand; only one on leading strand
6. DNA polymerase
  1. responsible for covalently linking deoxyribonucleotide monophosphate for the 3' end of the growing strand
  2. diff. kinds with diff. functions
    1. proof-reading
    2. polymerization
    3. removal or RNA primers once not needed
7. DNA ligase connects the gaps between the fragments
The direction of replication
1. DNA polymerase can only add nucleotides to the 3' end of the primer
2. begins at site called origin of replication
3. occurs in both directions away from origin
4. five carbons of deoxyribose sugar have a number
5. phosphate group added to 3' of the deoxyribose of the nucleotide at the end of the chain
6. replications occurs in a 5' to 3' direction
Non-coding regions of DNA have important functions
1. cellular machinery operates according to genetic code
2. coding sequeneces - DNA sequences used to produce polypeptides
3. used as guide to produce tRNA and rRNA
4. enhancers and silencers regulate gene expression
5. most genome non-coding in eukaryotes
6. repetitive sequences common
  1. moderately
  2. highly
    1. satellite DNA
  3. 5-60% (60% in humans)
  4. one occurs on the ends of eukaryotic chromosomes; telomere
    1. protects cells from losing genes that cannot replicate during interphase

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