Dna replication

• What is the dna structure in prokaryotes?

  Single circular chromosomes which are condensed in the nucleoid via DNA supercoiling, DNA interacts with cytoplasm and therefore transcription and translation occur simultaneously. Only one copy of each gene. efficient and compact
• When does replication occur?

  When cell is ready to divide
• What are the properties of replciation?

  Semiconservative process , starts at a single point and is bidrectional , dNTP , 5'-3
• Explain the semi conservative property?


  Each daughter molecule receives one parental strand and has a newly synthesized strand
• Explain the bidirectional characteristic?

  DNA replication starts in a single well-define point: OriC. Bidirectional: Two replication forks in which the DNA is actively synthesized
• Explain the activated nucleotides?

  Template dna: does not participate in the reaction but defines which substrate is going to be incorporated. In each reaction a dNTP is added to a polynucleotidic chain. PPI hydrolysis provides the Gibbs energy needed
• Explain the 5'-3' direction property?


  There is a leading and a lagging strand. The leading strand replicates from 5'-3'. However, due to the structure the lagging strand has to travel in opposite direction forming Okazaki fragments
• What are Okazaki fragments?


  Short lengths of single-stranded DNA made on the lagging strand.
• What are some proteins involved in replication?

  Topoisomerases: relieve the strain caused by DNA unwinding

  
  

  Replisome: Multiprotein molecular machinery responsible for replication of DNA

  
  

  Ligases: Join DNA strands

  
  

  DNA polymerase: polymerization activity

  Helicase: Unwind DNA

  primasas: Synthesize

  short RNA sequences

  SSP: Monomeric proteins that bind to single stranded DNA to prevent double strand formation
• What are the types of nuclease activity?

  Endonuclease , exonuclease (3'-5') and exonuclease (5'-3')
• What are the characteristics of DNA polymerase?

  Addition of a new nucleotide through a phosphodiester linkage to a 3OH group. AS a consequence DNa polymerase cannot start a new polynucleotidic strain , only adds nucleotides to pre-existing chains , requires a primer (short RNA sequence synthesized by an RNA polymerase (primase))

  Low rate of mistakes: Active site pairing , proofreading activity
• What is the clamp?

  A dimeric protein that increases processivity
• Steps in the replication?

  Initiation , elongation , termination
• What occurs during initiation?

  OriC presents three 13 bp sequences in the same orientation and four 9 bp sequences in different orientations. Proteins called DnaA recognize and bind the 9 bp repeats; a loop is formed. HU is an accessory factor to assist the action of DnaA opening the double helix. DnaB and DnaC bind to 13 bp repeats and unwind Dna. Initiation Bubble. Sequential recruitment of primase and DNA pol 3.
• What occurs in elongation?

  Dna polymerase 3 and DnaB helicase and the primase bind to form the replication fork. Gyrase eliminates the strain caused by DNA unwinding. DNA polymerase catalyses two 5'-3' syntheses in opposite directions. Dna in the lagging strand forms a loop, so the direction of DNA synthesis coincides with the movement of the fork. Lagging strand requires a new primer and new clamp for each Okazaki fragment.
• What is a nick?

  A point in which two nucleotides are not bound (DNA polymerase 3 cannot remove primers but moves it)
• How to fix a nick?

  DNA ligase catalyse the phosphodiester linkage without adding a nucleotide
• What occurs in termination?

  Ter sequences are termination sequences opposite to OriC. A topoisomerase releases the catenated chromosomes. DNA is methylated by Dam, allows mismatch repair of the unmethylated strand
• What are the differences between prokaryotic and eukaryotic replication?

  Dna polymerases with an identical mechanism of action: alpha (primase) , Beta (repair) (mitochondrial DNA) , delta. Each chromosome has many initiation points (OriC), a slower process due to presence of nucleosomes , shorter Okazaki fragments , DNA methylation in cytosines , problem of having linear DNA molecules: Telomerase
• What are the characteristics of telomerase?

  ends of linear chromosome cannot be replicated , 50 nucleotides lost each cell cycle (telomeres). Reverse transcriptase: RNA template to synthesize the TG strand
• What is the Hayflick limit?


  Normal cells have limited capacity to divide except immortal cell types and tumour cells