Replication Flashcards
Complementary DNA pairs
A+T and C+G
Pyrimadine
C+T
Purine
G+A
How many bp in a turn of the DNA double helix?
10 bp/turn
Length of a helical turn?
3.4 nm/turn
Distance between bases?
0.34 nm
DNA replication is?
Semi-conservative:
After one round of DNA replication, each DNA contains one parental and one newly-synthesized strand.
What is DNA synthesis not?
Conservative: Formation of a completely new molecule; after one round one parental molecule and one new molecule.
Dispersive: Parental molecule is completely disrupted and reassembled to make a new strand; half of the ds-DNA is the parental and the other half of the helix is new.
Why is DNA replication important?
Ensures an exact copy.
If DNA failed to copy, meiosis and mitosis would not occur, and life would not exist.
What are the three modes of DNA replication?
Theta replication, Rolling circle replication, Linear chromosome replication.
What is theta replication?
(Unwind, replication bubble, replication fork, replication)
- Occurs in most circular DNA
- Replication is bidirectional
- Two circular DNA molecules are produced
What is rolling circle replication?
(Break in nucleotide strand, replication of inner strand, cleavage releases a ss-linear DNA and ds-DNA)
- Occurs in the F factor of some viruses
- Replication is unidirectional
- Produces multiple circular DNA molecules
What is linear chromosome replication?
(Unwinding, replication bubble, DNA synthesis on both strands at each end of the bubble as replication forks form, forks meet and segments fuse)
- In eukaryotes
- Replication is bidirectional
- Produces two linear DNA molecules
What is required for DNA replication?
- Requires magnesium
- DNA polymerase
- dNTPs
- A template strand
- RNA primer
What are the features of DNA replication?
- Synthesized 5’ to 3’
- New strand is complementary and antiparallel to template
- Held together by hydrogen bonds
DNA chains are susceptible to nuclease cleavage…
… can cleave a phosphate group attached to 5’ or 3’ carbon
(gamma, beta, alpha phosphate attached to nitrogenous base)
What is a nucleotide?
Phosphate group + a sugar + a nitrogenous base
Where is DNA synthesis continuous?
On the leading strand
- By polymerase III
Where is DNA synthesis discontinuous?
On the lagging strand creating Okazaki fragments (short fragments of DNA produced by discontinuous synthesis).
- By polymerase III and polymerase I fills in the gaps
Where does DNA replication begin?
From an “origin”
What does prokaryotic DNA replication consist of?
- oriC orgin site
- four dnaA binding sites
- an AT rich region that undergoes strand seperatio n
What is helicase?
Unwinds DNA in the 5’ to 3’ direction
- Breaks hydrogen bonds and moves the replication fork
- Binds to the lagging-strand template at each replication fork
What are single-strand-binding proteins?
Immediately binds to the single-stranded DNA after helicase unwinds it.
- Keeps DNA linear so it does not fold back on itself
- Allows DNA to be easily replicable
What is DNA gyrase?
Topoisomerase II:
- Encircles DNA and removes torsional stress caused by the helicase unwinding which causes positive supercoils that can stop replicaiton (no more unwinding)
- Nicks DNA to release supercoils
- Relieves strain ahead of the replication fork
What is an initiator protein?
DnaA, binds to oriC and causes a short stretch of DNA to unwind so helicase and ssb proteins can attach.
What is topoisomerase II?
DNA gyrase
What is DNA primase?
Synthesizes a short RNA primer that provides the 3’ OH end for the DNA polymerase to begin DNA synthesis
- Essential to start replication
- Only one primer is needed on the leading strand
- A primer is needed for every Okazaki fragment
How many DNA polymerases are in E. coli?
Five: Polymerase I, II, II, IV, and V.
Which polymerases in prokaryotes are used for replication?
Polymerases I and III
Which polymerases in E. coli are used for DNA repair?
Polymerase II, IV, and V
What is the function of DNA polymerase I?
- Aids in the removal of primers
- Has 5’ to 3’ polymerase and 5’ to 3’ exonuclease activity Can remove RNA primer and re-synthesize a short tract of DNA
- Proofreading: has 3’ to 5’ exonuclease activity
- Short tract synthesis
- Mostly lagging strand
What is the function of DNA polymerase III?
- Main polymerase
- 5’ to 3’ polymerase activity
- Proofreading: has 3’ to 5’ exonuclease
- DOES NOT HAVE: 5’ to 3’ exonuclease activity; in this case polymerase I takes over
What is 5’ to 3’ exonuclease?
Can chew away at 5’ RNA primers and fill gaps with DNA.
What is 3’ to 5’ exonuclease?
Corrects mistakes polymerase may make
What is the beta-sliding clamp?
A ring-shaped polypeptide that encircles the DNA
- Interacts with polymerase III
- Helps to keep ssb proteins on
What is DNA ligase?
Seals the nick in fragments of DNA with a phosphodiester bond
How are both strands able to replicate simultaneously?
DNA must form a loop so that 5’ to 3’ synthesis can take place on both anti-parallel strands
In what phase of the eukaryotic cell cycle does replication begin?
Prepared for replication in G1 phase, actual replicating begins in the S phase.
What are short telomeres associated with?
Low telomerase activity:
Cellular senescence and cell death, diseases causing premature aging: Progeria, werner’s syndrom.
What are long telomeres associated with?
High telomerase activity: cancer (promotes growth)
What are unique aspects of eukaryotic chromosome replication?
- Shorter RNA primers and Okazaki fragments
- DNA replication only during S phase
- Multiple polymerases (15)
- Bidirectional replication from multiple origins and on each chromosome
- Nucleosomes
- Telomeres: shorten at each round of eukaryotic replication
What is the problem with telomeres?
Chromosome end will be degraded causing chromosome shortening during every round of replication.
Telomerase activity extends eukaryotic chromosome ends in replicating cells to prevent this.
What is the first step in the reaction sequence?
Proofread and correct mismatch pairs.
Which group of enzymes is responsible for making breaks in DNA?
Nucleases
What is the role of RNA primer?
To provide a free 3’OH substrate for polymerase III
