6 - DNA replication and mutations Flashcards
what phase of the cell cycle does DNA replication occur?
the S phase
what is chromatin?
DNA molecule and protein (histones) complex
what is a chromatid
one of two identical chromosomes joined together at their centromere
what are chromosomes?
complete densely packed chromatin structure
what do diploid organisms including most animals and cells have?
paired chromosomes
what do haploid organisms have?
only one set of chromosomes
what is produced by mitosis?
two identical diploid cells.
what is created in meiosis?
four haploid cells
what does replicated DNA consist of?
one strand of the old DNA (parent) and one strand of the new DNA (daughter)
DNA strands represent templated for DNA replication
what are the three proposed DNA replication models?
- semiconservative model
- conservative model
- dispersive model
what did the meselon-stahl experiment prove?
it disproved the conservative and dispersive models.
what are the three DNA replication stages?
initiation and unwinding
primer synthesis
elongation
where does DNA replication begin?
replication begins at specific DNA sequences known as the ‘origins or replication’ or ‘ori’
these create a ‘replication bubble’ in the DNA strand and replication happens in both directions.
in circular and linear DNA how many ori are there?
in circular DNA there is a single ori
in linear DNA there are multiple ori
…
these ori sequences are AT rich as the hydrogen bonds (2H bonds) between adenine and thymine are weaker than between cytosine and guanine (3H bonds)
what is a replicon?
eukaryotic chromosomes contain numerous replication origins, allowing for DNA synthesis to proceed in parallel at many sites simultaneously.
each pair of replication forks is referred to as a “replicon”
how can replicons be visualised?
replicons can be visualised as bubbles of replication DNA that expand in both directions.
eventually, all replicons merge into a single large bubble, until replication terminates at the telomeres.
how many replication forks does each replication have?
each replication has two replication forks, moving in opposite directions.
ultimately, replication forks meet, until replication of each template strand is complete.
how many base pairs apart are ori in eukaryotes?
ori occur about 40,00 base pairs apart allowing each chromosome to be replicated very fast.
how many nucleotides per second are replicated in human?
50 nucleotides per second in human
how long does it take for complete replication of genome in human cells?
can take 5-10 hours.
replication dosen’t happen to all chromosomes at the sae time.
how fast can ori copy in prokaryotes?
Prokaryotes don’t have multiple Ori, however they can copy at speeds of >1000 nucleotides/second
what direction does DNA polymerase copy in?
5’ to 3’ direction so one strand is continuous (leading) and the other is lagging
what is DNA helicase?
the enzyme that separates the two strands during replication
what is primase?
enzyme that synthesises short nucleotide sequences called primers. RNA primers serve as a starting point for DNA synthesis.
what are single-stand binding proteins?
bind to and stabilise single-stranded DNA
what is DNA ligase?
joins fragments of DNA together
what is topoisomerase
untangles the DNA strand which becomes supercoiled
how does topoisomerase untangle the DNA?
it untangles the DNA strand near the replication fork by cutting up one or both of the strand, straightening out the coils and re-joining the strands.
how does topoisomerase effect the strands?
cut and uncut strands und up having the same chemical composition cut different topology (isomer)
what do DNA polymerase do?
catalyses the addition of complementary nucleotides to the growing chin and catalyses phosphodiester bonds between nucleotides.
can DNA polymerase catalyse a new strand from scratch?
DNA polymerase cannot catalyse a new strand from scratch - it needs an existing strand from scratch to elongate from the 3’ -OH end.
so an RNA primer is necessary to initiate DNA replication
what are the different types of DNA polymerase?
numbered ones are found in prokaryotes (e.g. Pol I, II, III) and lettered ones are found in eukaryotes (e.g. Pol α, δ, ε).
what direction is the template for the leading strand orientated in?
3’ to 5’ allowing continuous replication in the 5’ to 3’ direction.
what steps are involved in the leading strand?
- primase, and RNA polymerase makes a short strand of RNA known as a primer, which pairs to the 3’ end of the leading strand of DNA. this acts as a starting point for DNA replication
- DNA polymerase elongates the new strand by adding nucleotides to their complementary base pairs and catalyses phosphodiester bonds between the nucleotides.
- replication continues until the 5’ end of the template has been reached
- the primer then breaks off at the 3’ end of the template and polymerase activity is used to replace the RNA primer with DNA nucleotides.
how is the lagging strand replicated?
in the 5’ to 3’ direction, so the the lagging strand needs to be copied in short fragments = discontinuous replication.
what are the short fragments in the lagging strand called?
Okazaki fragments
what are the lagging strand fragments joined up by?
DNA ligase
what are the steps of replicating the lagging strand?
- RNA primer binds to site on lagging strand
- Okazaki fragment formed along a 5’ to 3’ direction by DNA polymerase
- another primer binds to site behind the first okazaki fragment
- another fragment produced
- ribonuclease activity removes the RNA primers
- gaps then filler in with DNA by polymerase activity
- DNA ligase joins the fragments.
how common are errors in RNA during transcription and translation?
Errors in RNA during transcription and translation are about 1 in 104-105. however, an error in a mRNA strand will not have lasting damaging effects.
what happens with errors in DNA replication?
they are passed onto the next generation
what does proof-reading do?
acts as a first line of defence in correcting errors caused by polymerase.
how does DNA polymerase proof read?
DNA polymerase proof-reads as it synthesises the growing DNA strand by double checking the correct nucleotide has been added before it catalyses the addition of the nucleotide.
The enzyme undergoes a conformal change before it adds the next nucleotide to the growing chain. An incorrect nucleotide will likely dissociate at this step.
what direction does DNA polymerase proof-read?
in the 3’ to 5’ direction
Exonuclease activity: as new nucleotides are added to the replicated strand, it scans along behind
what does DNA mis-match repair do?
Addresses incorrect pairing of parent nucleotide to daughter nucleotide missed from proof-reading and exonuclease activity.
what are mis-matches caused by?
Mis-matches were initially thought to be caused by tautomerization- slightly different bonding patterns on nucleotides caused by a shift in their form. This leads to A pairing with C, and T pairing with G. Nowadays believed to be caused by a wobble
what are mis-match repair proteins called?
Mis-match repair proteins called ‘Mut’ proteins act as the next line of defence in DNA replication errors
what are tautomers?
Tautomer’s are isomers with different bonding patterns (e.g. position of proton)
what are the two types of DNA mutations?
Endogenous and exogenous
what is an endogenous mutation?
mutations resulting from cellular processes (errors during DNA replication, chemical modifications)
what is an exogenous mutation?
mutation stemming from external causes e.g. environmental factors (smoking, sunlight, radiation, external mutagens)
what is a silent mutation?
does not change the protein it codes for
what is a nonsense mutation?
results in a premature stop codon. resulting protein is non-functional
what is a missense mutation?
a different amino acid is produced, changing the resulting protein.
what is a frameshift mutation?
caused by an insertion or deletion of one or two base pairs which shifts the way the sequence is read: provided its not a multiple of three
what is an example of a point mutation
sickle cell disease.
