L2- DNA replication

Overall, what happens in interphase and mitotic phase?

Interphase (G1,S,G2)- cell grows and makes a copy of it’s DNA

Mitotic phase- Cell separates its DNA and forms two new cells

What is semi-conservative replication?

DNA acts as a template for it’s own replication

Which enzyme unwinds the DNA?

DNA helicase

Which enzyme synthesis DNA in the 5’-3’ direction?

DNA polymerase

Which enzyme relieves the tension and twisting in the DNA?

DNA topoisomerase

Which enzyme synthesis RNA primers?

DNA primase

Whcih enzyme degrades RNA primers?

Ribonucleases

Which enzyme joins DNA fragments?

DNA ligase

Which enzyme replicates the ends of the chromosome?

Telomerase

Which way does DNA replication occur?

5’-3’ direction

On which strand is replication continuous?

The leading strand

On which strand is replication discontinuous, and why is this?

The lagging strand- replication cannot progress in the opposite direction, so it must wait form some DNA to unwind

What are the small fragments called produced on the lagging strand?

Okazaki fragments

What is the replication fork?

Occurs when helicase unzips the DNA double helix, and the separated strands [provide a template for new strands

What is a sliding clamp?

DNA polymerase remains attached to the DNA template by interaction with a sliding clamp (protein). A new clamp has to be loaded on the lagging strand as each okazaki fragment is synthesised

On the lagging strand, after the primers are removed what happens next?

DNA polymerase fills in the gaps

DNA ligase joins the fragments together

Why is a primer needed?

To provide a 3’ OH group for the nucleotides to be added to

What is a telomere?

Repetitive regions at the ends of chromosomes, acting as caps to protect the internal regions of chromosomes (worn down a small amount in each round of replication)

TTAGGG repeated hundred/ thousands of times

How does the telomere shorten after each round of replication?

• In lagging strand, okazaki fragments cannot cover the end of the chromosome (when final RNA primer is removed, a gap is left rather than being replaced by DNA)

• This part gets uncopied in each round of replication, leaving a single-stranded overhang

What is the solution to the telomere shortening after every round of DNA replication?

Some cells have telomerase- an enzyme that can make DNA using RNA as a template

• Binds to RNA molecule containing sequence complementary to telomeric repeat

• When overhang is long enough, a matching strand can be made by DNA polymerase a

How can polymerase fix a mistake by proofreading, if the incorrect nucleotide is added on?

Polymerase detects the bases are incorrect, and goes back in the 3’-5’ direction, using exonuclease to remove the incorrect base. It will then continue in the 5’-3’ direction

What can cause DNA to be damaged?

• UV light

• ionising radiation exposure

• Toxic chemical agents

• Reactive oxygen species

What could happen as a result of DNA damage?

Mutations in genes, leading to altered coding for proteins, resulting in loss or gain of function

Cancer

What is depurination?

Occurs as a result of damage to the DNA, involving the removal of purines (guanine or adenine)

What is deamination?

Occurs as a result of damage to the DNA, involving a C to U transition

What is a thymidine dimer?

When two adjacent thymine bases become covalently attached due to exposure to environmental factors, damaging the DNA

What is a double-stranded break?

Some environmental factors (e.g. high energy radiation) can cause chromosomes to literally split in two.

Depending on where the break is, large number of bases and encoded information can be lost

What are the two classes of mutation?

• Gain of function- leads to increased or alternative activity

• Loss of function- leads to decreased activity

Substituting a nucleotide can alter the codon in which three ways?

1. Silent

2. Missense

3. Nonsense

How does a silent mutation occur?

When a nucleotide substitution results in a different codon, that still encodes for the same amino acid

Protein is unaffected

What is a missense mutation and how does it occur?

When a nucleotide substitution results in a codon encoding for a different amino acid

Primary protein sequence is altered

What possible consequences of a missense mutation are there?

1. Conservative substitution- similar amino acid R group, in size and charge. Similar protein shape and function

2. Radical substitution- New amino acid R group has a very different size or charge, protein has an altered secondary or tertiary structure, affecting function

What is a nonsense mutation?

When a nucleotide substitution results in a stop codon, stopping translation

Protein is truncated and may not function properly

What is an insertion mutation?

Adding a nucleotide into the sequence, resulting in a frameshift and all codons downstream are affected

What is a deletion mutation?

A nucleotide is removed, resulting in a frameshift and all codons downstream are affected