DNA replication

• DNA replication

  • The cells that make up multicellular organisms are derived from existing cells by the process of division. Cell division occurs in two main stages:
  • Nuclear division - when the nucleus divides - mitosis and meiosis
  • Cytokinesis - when the whole cell divides
  • What has to happen before the nucleus divides?
  • DNA replication
• DNA replication

  Any substance acting as genetic material must be capable of:

  • Encoding information - DNA uses a sequence of letters
  • Exact replication - all of the information must be precisely copied every time a cell divides. This happens because of the complementary base pairing rule
  • An error in the copying of the code would be classified as a mutation
• DNA replication

  • The specific complementary base-pairing in double-stranded DNA provides an obvious mechanism for its replication - How?
  • All that need happen is for the two strands to separate, then each strand acts as a precise template for the synthesis of its partner.
  • Wherever one strand has A, its partner must have T; 
  • Wherever one strand has C, its partner must have G.
• Semi-conservative replication

  1. The 4 nucleotides with their bases must be present
  2. Both strands of the DNA molecule act as a template for the attachment of the nucleotides
  3. The enzyme DNA polymerase
  4. A source of energy is required
• DNA Polymerase

  • DNA polymerase is a doughnut-shaped molecule made of ten separate polypeptides.
  • Its role is to join free nucleotides by condensation reactions to form phosphodiester bonds and new polynucleotide strands
  • DNA polymerase also proof reads the new strand to ensure there are no errors (mutations)
• DNA REP PROCESS

  • DNA helicase breaks the hydrogen bonds linking the base pairs of DNA
  • The double helix separates into its two strands
  • Each exposed polynucleotide strand then acts as a template to which complementary free nucleotides bind by specific base pairing
  • Nucleotides are joined in a condensation reaction by the enzyme DNA polymerase to form the missing polynucleotide strand on each of the two original polynucleotide strands of DNA
  • Each of the new DNA molecules contains one of the original DNA strands, that is, half the original DNA has been saved and built into each new DNA molecule. 
• DNA Replication

  The original strand is called the template strand. In DNA replication there will be 2 template strands. The new strands are called the newly synthesised strands.
• Mechanism of Replication: Summary  

  1. The double helix is unwound/unzipped by a complex of enzymes including DNA helicase which breaks the hydrogen bonds between the base pairs This allows both of the strands to now act as templates.
  2. The enzyme DNA polymerase synthesises a new 5’-3’ strand by inserting complementary nucleotides one at a time. 
  3. The nucleotides are inserted as nucleotide triphosphates: two phosphate groups are released as each is bonded into place. The nucleotides have been ‘activated’ by the phosphate groups from ATP.
• In which direction can DNA polymerase build a new strand?

  5’ to 3’ direction
• How does DNA polymerase attach to the original strand during replication?

  It attaches to the 3’ end of the original strand
• What happens to DNA as it is unzipped during replication?

  DNA is unzipped from 3’ towards 5’ end
• What is the leading strand in DNA replication?

  The leading strand is synthesised continuously
• What is the name of the strand that is created opposite the leading strand?

  Lagging strand
• How does DNA polymerase move on the lagging strand?

  It moves away from the replication fork
• What direction does DNA polymerase move on the lagging strand?

  From 5’ to 3’ end
• What are Okazaki fragments?

  Short segments of lagging DNA strand
• What role does DNA ligase play in DNA replication?

  It joins the lagging strand segments together
• What are the key differences between leading and lagging strands in DNA replication?

  • Leading strand: synthesized continuously
  • Lagging strand: synthesized in short Okazaki fragments
  • Leading strand: moves towards replication fork
  • Lagging strand: moves away from replication fork
• DNA Replication

  • DNA polymerase can therefore only make sequences from 5’-3’ in small sections, because the enzyme is moving away from the fork, so needs to release from the DNA and reattach at different points.
  • These small sections are called Okazaki fragments, which are synthesised in a discontinuous manner. The lagging strand needs a new primer for the synthesis of each Okazaki fragment. DNA polymerase can then add a short row of bases in 5’-3’ direction. 
  • The next primer is added further down the lagging strand, another fragment is made, and the process is repeated.
• What direction do DNA strands in a double helix run?

  Opposite directions
• Why is the direction of DNA strands a problem for replication?

  DNA polymerase can only add bases 5’-3’
• How does the leading strand differ from the lagging strand during replication?

  The leading strand is made continuously
• What does the replication fork do during DNA replication?

  It unravels the DNA ahead
• How does DNA polymerase add bases to the leading strand?

  In a 5’-3’ direction
• Why can't the lagging strand be made continuously by DNA polymerase?

  It runs in the opposite direction
• What are the mechanisms used to copy the leading and lagging strands?

  Leading strand is continuous; lagging strand is not
• DNA Replication

  • DNA replication goes in the 5' to 3' direction because DNA polymerase acts on the 3'-OH of the existing strand for adding free nucleotides.
  • Nucleotides cannot be added to the phosphate (5’) end because of the shape of the active site of the enzyme DNA polymerase which would not be the complementary shape.