Macromolecular Synthesis-DNA Replication Flashcards
Characteristics of DNA replication (3)
- bacterial genomes are circular
- DNA synthesis is semi-conservative
- proved by Meselson and Stahl experiment
Fidelity in DNA replication
- base selection
- 3’ to 5’ exonuclease
- mismatch repair
- involved multiple DNA pol.
Key Points of DNA rep. (5)
- DNA pol. add nucleotides to 3’ end
- DNA pol can only extend existing nucleic acid chains (can’t make new ones)
- DNA synthesis begins at the origin and proceeds bidirectionally
- DNA synthesis is continuous on leading strand
- lagging strand is made in pieces that are ligated together
Key Players of Replication-Primase (DnaG)
- RNA pol
- makes RNA primer
Key Players of Replication-Polymerase III
- DNA pol
- the workhorse pol
- has proofreading activity
Key Players of Replication-SSB
- single stranding binding proteins
- binds DNA to maintain in ss form
Key Players of Replication-Polymerase I
-replaces RNA primer ribonucleotides w/ deoxyribonucleotides
Early Steps in Replication-formation of open complex (2)
- DnaA binds specific sequences in OriC (w/help from histone like protein HU
- DNA unwinds (strands separate) in an ATP-dependent manner
- SSBs bind to keep open complex open
Early Steps in Replication-Prepriming complex (2)
- DnaB (helicase) binds to both ends of open complex
2. gives 2 replication forks going in opposite directions
Key Players of Replication-DNA ligase
-links okazaki fragments together
Key Players of Replication-Helicase (DnaB)
-unwinds the DNA ahead of the replication fork
Key Players of Replication-Tus
- binds Ter
- blocks replisome until the other one shows up
Key Players of Replication-Replisome
-complex of proteins that make up the replication fork
Key Players of Replication-HU
histone-like protein that helps DnaA bind the ori
Priming (4)
- Primase binds and makes 12 nt piece of RNA
- DNA pol III can initiate DNA synthesis, adding deoxyribonucleic acids one at a time to the 3’ end
- extension occurs continuously on the leading strand
- synthesis of lagging strand is made in pieces called okazaki fragments
Okazaki Fragments (5)
- same steps as for synthesis of leading strand
- usually 1000 bp
- Pol III stops when it reaches last RNA primer
- RNA primer is removed by Pol I (5’ to 3’ exonuclease activity)
- break sealed by DNA ligase
Termination
- Tus binds Ter
- Ter allows one way replication
- Ter stops replisome, thought to stop the helices DnaB
- other replisome shows up and dislodges Tus
- replication is done
- chromosomes end up tangled, resolved by recombination or decantation by Topo IV
Chromosome Dimer formation and resolution
- chromosomes get tangled during replication and form dimers
- dimers resolved by site specific recombination at dif sites
- the dif sites are lined up by FtsK, which is localized to the septum
- XerC/D is activated-the recombinase that acts on dif sites
DNA Supercoiling (5)
- DNA duplex is a right handed helix
- 10.5 bases/ turn
- overwinding=positive supercoiling, Underwinding=negative supercoiling
- supercoiling is energy stored in DNA molecules as twist/writhe
- superhelical tension can be used to aid processes (packaging, open complex formation)
Topoisomerases (3)
- enzymes that introduce or relieve supercoils by breaking the phosphate bond of DNA
1. topo I removes supercoils
2. Gyrase-introduces supercoils
3. topo IV-removes catenations
Histone like proteins
- compact DNA
- small and abundant
- some are site specific and some bind in a more general way
- can serve as accessory factors to help initiate processes (replication, recombination)
- can be general repressors (e.g. HNS can repress pol binding by occupying space on DNA)
