- Helicase unwinds the parental double helix at the origin of replication→ forms a Y-shaped replication fork
- Origin of replication: short, stretch of DNA with a specific sequence of nucleotides
- Single-stranded binding protein attaches to each strand of uncoiled DNA to keep separate
- As helicase unwinds the DNA → forces the double helix to twist → group of enzymes topoisomerases break and rejoin the double helix → allow twists to unravel and prevent knots
- Primase: an enzyme that initiates DNA replication at the origins of replication by placing an initial, short RNA nucleotide strand (RNA primer) using parental DNA as a template
- Primase also slows replication fork
- Need primase and primers bcuz DNA Polymerase can only attach to 3’ end of an already existing complementary strand → belongs elongation of new DNA at replication fork by assembling new (complementary) strand in the antiparallel 5’ → 3’ direction
- Leading strand and every Okazaki fragment on lagging strand must begin with primer
- Since DNA consists of two opposing DNA strands, uncoiled DNA consists of 3’ → 5’ template strand and 5’ → 3’ template strand
- Leading strand: For the 3’ → 5’ strand, replication occurs continuously DNA polymerase moves towards the replication fork
- Lagging Strand: For the 5’ → 3’ template strand, replication occurs discontinuously as DNA polymerase moves away from the uncoiling replication fork
This is bcuz it can assemble nucleotides only as it moves in 5’ → 3’ direction; takes more time to assemble
- After each complementary segment is assembled & DNA pol III reaches next RNA primer it must return back to the replication fork to begin assembling the next segments
- Okazaki fragments: Short segments of complementary DNA; have 5′ RNA nucleotides & DNA nucleotides 3′
- RNA nucleotides of RNA Primer are later replaced with DNA by DNA pol 1
- DNA ligase joins the sugar-phosphate backbones of Okazaki fragments and closes up gaps thru covalent bonds
Antiparallel Elongation
- Energy for elongation provided by two additional phosphates that are attached to each nucleotide (total of 3 attached to nitrogen base)
Breaking the bonds that holds extra two provides chemical energy for process
Replication of Telomeres
Big problem when replication reaches the end of DNA strand
- Eukaryotes can’t complete 5’ end of lagging strand bcuz last primer removed and no 3’ end for DNA pol to add DNA
- Result: DNA loss → shorter and shorter DNA molecules with uneven ends → can trigger apoptosis
- To solve this problem telomerase attaches to the end of the template strand and extends template by adding telomeres
- Telomeres: noncoding, special DNA sequence, 5’-TTAGGG-3
- Telomere Functions: Allows elongation of lagging strand to continue, stops staggered ends, and postpones loss of DNA in replicated chromosomes
- Telomerase activity declines as cells age → once stops, chromosome becomes shorter with each replication → eventually important DNA at end of the chromosome is lost → nonfunctional daughter cells
- Telomerase activity high in cancer cells and tumors → prolongs life
DNA Proofreading and Repair
- DNA replication error low bcuz of base pairings and DNA polymerase but not perfect so cells have mechanisms to repair errors
- w/o mechanisms → accumulate cancer-causing errors
- Proofreading: Polymerase proofreads newly made DNA, replacing any incorrect nucleotides with the correct ones
- Mismatch Repair Proteins: other enzymes correct errors in base pairing
- Excision repair proteins: nucleases identify & cut out damaged DNA → DNA pol 1 replaces → DNA ligase joins
Error rate after proofreading is low but not 0
- Mutations may be passed onto next generations
- Leads to genetic variation ⇒ natural selection