DNA Replication Flashcards
DNA replication is ____ and ______
Bidirectional (goes either way from origin of rep)
Semiconservative (one conserved parental strand, one newly synth daughter strand)
DNA synthesis ALWAYS proceeds in _____ direction
5’ to 3’
Leading strand has ______ replication, lagging strand has ______replication
continuous
discontinuous
origin of replication
AT rich sequences recognized by origin binding proteins
Bacterial: 1
Eukaryotic: multiple
DNA polymerase does what?
Synthesizes DNA strand 5’ to 3’ direction.
Replication forks
Site where DNA synthesis is occurring.
Origin binding proteins
They recognize and bind to origin of replication (AT rich sequences)
Helix unwinds _____ of replication fork by _____
ahead
helicases
What can hold DNA in single stranded conformation?
single-strand binding proteins
(Protects ssDNA from nucleases)
RPA: humans
SSB: bacteria
What problems must be overcome by DNA Pol in order to copy DNA?
- Unwinding (can’t melt strands to separate them)
- Primer: DNA Pol CANNOT initiate a new strand, it can only elongate an existing primer.
- Polarity (Strands have opposite polarity, but DNA Pol starts at 3’ end to make strands 5’-3’)
topoisomerase
Moves ahead of replication fork to prevent supercoiling by breaking and rejoining DNA backbone.
DNA gyrase is in prokaryotes (inhibited by quinolones)
Topo II makes double stranded breaks
Characteristics of replication origins
- unique segments w/ multiple short repeats
- rich in AT base pairs
- recognized by multimeric origin binding proteins
primase
enzyme that catalyzes a reaction that copies the parental strand to form RNA primer (which has about 10 nucleosides)
DNA polymerase
Adds deoxyribonucleotides to 3’ hydroxyls of RNA primer and to ends of growing DNA strand
Two DNA polymerases that carry out DNA rep?
DNA Pol I
DNA Pol III
DNA Pol I
- “clean up” in DNA replic/repair.(3’ to 5’ exonuclease activity for proofreading)
- Also removes RNA primer from DNA by using 5’ to 3’ exonuclease activity
- 5’ to 3’ DNA Pol activity (needs 3’ OH primer and DNA template strand)
DNA Pol III
MAJOR replicative enzyme b/c it has clamp which keeps it attached to DNA over long dist. High processivity.
DNA ligase
JOINS Okazaki fragments, catalyzes formation of phosphodiester bonds b/t 3’ OH and 5’ phosphate group b/t two polynucleotide chains.
Cofactor for ligase in eukaryotes and prokaryotes?
ATP for euk
NAD+ for pro
Sliding clamp for euk and prok
PCNA for eukaryotic DNA Pol δ
Beta subunit for prokaryotic DNA pol III
(Clamp increases processivity of DNA pol)
Leading and lagging strands are synthesized concurrently via ______
looping of the lagging strand (solves polarity issue)
What does DNA Pol use to increase accuracy?
Discrim b/t correct and incorrect nucleotides by:
-ability to form hydrogen bonds (specifies correct pairing b/t complimentary bases
-Common geometry (AT, CG required to fit in active site of polymerase)
10^4-10^5 correct additions
Proofreading by DNA pol
increases accuracy to correct additions for every million to 100 million nucleotides
3’ to 5’ exonuclease activity of DNA Pol I AND DNA Pol III
Sliding Clamp on Euk and Prok
Prok: Beta clamp
Euk: Lagging strand: delta clamp, leading strand E (epsilon) clamp
Retroviral replication require conversion of _____ to ____ by _____
ssRNA to dsDNA by reverse transcriptase (reverse transcription)
Telomerase
RNA-dependent DNA Pol that maintains chromosomal ends by copying telomeric repeat sequences from RNA template.
- Repressed in normal cells
- De-repressed in cancer cells (and stem cells), normal cell death is NOT triggered–>tumor growth
End replication problem refers to which strand?
Lagging, because it needs RNA primer to begin synth of each lagging strand. But @ end, there is nothing for it to attach to. Last section can’t be synthesized. Telomeres get shorter and shorter.
