MIC102 Lecture 8 - S7

How is the initiation step regulated during DNA synthesis?

• cell must have enough ATP to initiate DNA replication

• two identical copies fo chromosome

• Enough DnaA must be enough to bind all DnaA boxes at replicative origin (oriC)

• and bound by DnaA ATP

SeqA

• GATC sequences scattered around oriC, methylase recognize the “A”, if initiation occurs, replicated starnds not yet methylated → SeqA high affinituy for hemimehtyalted GATC and hide priC DNA

Components of DNA

Deoxyribonucleotide (dNTP or nt)

• phosphate + deoxyribose + nitrogenous base

Backbone

• phosphate and deoxy ribose linedk via phosphodiester bond

Base Pairing

• G-C

• A-T

• Pyrimidines (one ring) - T,C

• Purines (double ring) - G,A

Antiparallel

Nt added to 3’ end to 5’ to 3’ direction

• DNA strand + CTP → DNA-CMP + PPi

What are some features fo DNA replication?

• semi-conservative (one parent and one daughter), fast and accurate

• Fast - After initiation - DNA replicated at 1000nt/s at each fork

• Accurate

  • 1 mismatched base / 100k

  • After proofreading and mismatch repair: 1 mismatch/ 10 bilion

Organize steps of initiation (5 steps)

1) At high [DnaA-ATP], Dna-ATP binds to binding sites at teh origin and break the hydrogen bonds holding strands together

2) Single-strand binding proteins keep the DNA strand apart

3) Helicase breaks H-bonds holding the double-stranded DNA together, separating the strands

4) Primase (RNAP) synthesizes a short RNA primer on both strands

5) full replisome assembles onto the replication bubble