DNA, the cell cycle and mitosis Flashcards
What is the role of DNA helicase?
DNA helicase unwinds double helix - ATP used to break h-bonds (b/c very stable molecule)
What does DNA polymerases require?
- Template strand (cannot start new chain from scratch)
- Oligonucleotide primer
- Deoxynucleotide triphosphates (dNTPs)
What direction does synthesis occur?
DNA synthesis in 5’-3’ direction (of strand being synthesized)
- Requires free 3’ hydroxyl (OH)
What drives the reaction?
- energy released by hydrolysis of triphosphates
What is the replication fork?
Reads 3’-5’
Places nucleotides 5’-3’
- site of DNA synthesis, fork moves along during process
- Daughter strand templates have opposite orientations, so fork not symmetrical
- Opens up as new bases are exposed
What is the origin of replication?
- discrete point where replication begins
How does the replication fork work?
Leading strand: 3’ end closest to replication fork - synthesis continuous (only need RNA primer once at beginning of strand)
Lagging strand: 3' end on opposite side - synthesis discontinuous in Okazaki fragments (copied backwards)
- RNA primes DNA for synthesis. DNA primase synthesizes short RNA fragment to be removes later - attach to 3’ of lagging strand
- DNA polymerase adds to 3’ of RNA primer and continues to synthesize Okazakis until reaches previous okazaki
- RNA primer of previous fragment removed by special ribonuclease using 5′ to 3′ exonuclease activity and replaced with DNA by repair DNA polymerase
- DNA ligase joins adjacent fragments to growing chain
- Synthesized chain 5’-3’, antiparallel to antisense strand
What other enzymes are involved?
Sliding clamp: forms ring around DNA to ensure DNA polymerase doesn’t fall off DNA strand
Single strand DNA binding protein: once DNA strands separated prevents them from locally folding
Explain semi conservative replication:
Each daughter cell inherits one old and one new strand, strands are complementary so each strand serves as template for next
- Automatically know the sequence of daughter strand
How do we avoid making mistakes?
Before nucleotide added, previous one checked for correct base pairing
- Incorrect bases removed by 3’-5’ exonuclease activity of DNA polymerase, then new one added
- phosphodiester bonds are hydrolysed prior to the replacement of incorrect nucleotides with the correct nucleotides
RNA Primers: Inaccurate RNA primers are replaced by accurate DNA.
How does replication occur in mammals?
- Large, linear DNA and multiple origins of replication at 100 kbp
- Each origin gives birdirectional forks
- Done when all forks meet
How does replication occur in e.coli:
- begins at unique origin; OriC
- Two replication forks proceed simultaneously in opposite directions (left and right) - bidirectional replication
- Two forks meet at other side of chromosome - replication then complete
What are the different phases of the cell cycle?
G1 (Gap1): each Chr present as single linear double helix; 10h
S: DNA synthesis; 9h
G2: each Chr has 2 identical sister chromatids: 4h
G0: cells stopped dividing
M: mitosis, chromatids separate into daughter cells; 1h
What are the different phases of mitosis?
Interphase (G2): Chr not visible
Late prophase: condense Chr
Metaphase: aligned on equator of spindle, microtubles attach to chromosomes
Anaphase: sister chromatids separate and pulled to pole
Telophase: move to opposite poles
Cytokenesis: 2 daughter cells
Interphase G2: unravel
