DNA replication Flashcards
what is the function of DNA helicase?
unwinds DNA
what is the function of DNA polymerase?
synthesis of DNA in 5’-3’ direction
what is the function of DNA topoisomerase?
relieves the tension in DNA
What is the function of DNA primase?
Synthesises RNA primers
What is the function of ribonuclease?
Degrades RNA primers
What is the function of DNA ligase?
Joins DNA fragments
How does DNA unwind?
- DNA helicase unwinds the DNA and uses ATP to proper itself along the DNA
- Single-strand DNA binding protein (SSB) binds and keeps the strands apart
- DNA topoisomerase relieves the tension
what is a primer?
short segment of RNA complementary to the template
what does DNA polymerase require?
template and primer
what does DNA synthesis take place in?
replication fork
What is the difference between replication on the leading and lagging strands?
Replication progresses 5’-3’ so it is continuous on the leading strand but it cannot progress in the opposite direction so replication is discontinuous on the lagging strand
What are the short DNA sequences synthesised on the lagging strand called?
Okazaki fragments
Explain the mechanism of the sliding clamp
- the DNA polymerase remains attached to the DNA template by interaction with a protein called sliding clamp
- a new clamp has to be loaded on the lagging strand as each Okazaki fragment is synthesised
- only one clamp is required on the leading strand
- clamp attached to replication fork
describe replication on the lagging strand
DNA primase attaches RNA to template
DNA polymerase III adds nucleotides until it reaches the previous primer
Ribonuclease H digests the RNA primer, leaving a gap
DNA polymerase I fills the gap
DNA ligase joins fragments together
what are telomeres?
Repetitive regions at the ends of chromosomes
G-rich series of repeat bases (TTAGGG repeated hundreds/thousands times in mammals)
What is the function of a telomere?
They act as caps to protect internal regions of chromosomes and are worn slightly each replication
Okazaki fragments can’t cover end of chromosome as primer would fall off end so no way of starting
explain the role of telomerase
- Telomerase are an RNA- dependent DNA polymerase, meaning an enzyme that can make DNA using RNA as a template
- the enzyme binds to a special RNA molecule that contains the sequence complementary to the telomeric repeat (AAUUCCC-TTAGGG)
- Telomerase recognises tip of existing repeat sequence, uses RNA template within enzyme to add additional repeats to the telomere DNA
- when the overhang is long enough, a matching strand can be made by DNA polymerase a, which has its own primase subunit
How often does DNA polymerase make a mistake?
Once every 10 000 000 pairs
What is the function of exonuclease?
Cutting out the wrong bases during proof-reading
Name 4 factors that can damage DNA
Ionising radiation exposure
UV light
Toxic chemical agents
Reactive oxygen species
What is depurination?
Removal of purine bases (guanine/adenine) leaving sugar-phosphate group. No DNA breaks
What is deamination?
No DNA breaks and results in C to U transition
What is a thymine dimer?
How do they arise?
What can they result in?
2 adjacent thymine bases become covalently attached to each other
Arise from UV light exposure and leads to stalling of replication machinery
Failure to repair thymine dimer is problem in xeroderma pigmentosum
what are the 3 types of mutations?
Point mutations
Insertion mutations
Deletion mutations
explain the 3 types of point mutation
A silent mutation results in a codon that still encodes same amino acid so the protein is unaffected and the organism’s phenotype is not significantly altered
A missense mutation results in a codon that encodes a different amino acid so the primary protein sequence is altered. can be conservative or radical:
Conservative when the new amino acid has similar function to the original e.g. similar R group size/charge and similar protein shape/function
Radical when new amino acid functions differently to original as the R group is different in charge/size so the protein may have altered secondary/tertiary structure and function is affected
A nonsense mutation results in a stop codon so the protein is truncated and may not function properly or at all
Explain insertion mutations
It changes the number of nucleotides so each one is shifted alone by one
Every codon from the point of insertion is different
Different amino acids are added to the protein and it is unlikely to function properly if at all
ORF has been moved, known as a FRAMESHIFT mutation
Explain deletion mutations
It removes a piece of DNA so each nucleotide is shifted alone by one
Every codon from the point of deletion is different
Different amino acids are added to the protein and it is unlikely to function properly if at all
ORF has been moved, known as a FRAMESHIFT mutation
List the 3 factors affecting tolerance of insertion/deletion mutations
Size: effect on protein structure
Location: introns/exons, coding potential and gene regulation
Open reading frame
what are the 2 classes of mutation?
gain of function
loss of function
what happens with a gain of function mutation? Give an example
DNA sequence changes that leads to an increase or alternative activity
e.g overactivity of a gene oviduct overrides existing control mechanism leading to cancerous cell
what happens with loss of function mutations. Give an example
a DNA sequence change that leads to decreased activity
e.g the nucleotide change leads to a loss of expression of the protein (null mutation)
what is the mechanisms for DNA repair
- Excision:
recognition and removal of damage using exonuclease - Repair
Re- synthesis of missing DNA using DNA polymerase - Joining
Sealing the nick using DNA ligase
How are thymidine dimers repaired?
Bubble formed, mistake cut out and gap repaired
Explain non-homologous end joining to repair double-stranded breaks
End ‘polished’ to produce blunt ends
Ends joined by DNA ligase
Results in loss of nucleotides and consequence depends on location
Very quick - useful for rapidly dividing cells
Explain homologous end joining to repair double-stranded breaks
End ‘polished’ to produce blunt ends
Complete repair so no sequence lost
Recombination between 2 corresponding regions of alleles
Other allele used as template
Slower - less useful for rapidly dividing cells but more accurate
Potential for introducing recessive traits in heterozygous individuals
mismatch repair reduces DNA replication mistakes to what?
1 in 10^9
