Proteins and enzymes of DNA replication - Week 2 Flashcards
what is the function of DNA polymerase
Matches the correct nucleotide and then joins adjacent nucleotides together
what is helicase
DNA unwinders to allow polymerase access
what is the function of single-strand binding proteins (SSB)
it prevents the strands of parental DNA from reforming a double helix
what do topoisomerases/gyrases do
they alter DNA conformation and relax the tension in the DNA strands
what do Ligase do
they join up DNA ends after synthesis
what happens in repair synthesis
repair DNA synthesis occurs during DNA replication.
If 1 strand of DNA is damaged, then that portion of the DNA is removed and new DNA material is synthesised to replace the portion of the fragment that has been removed. The enzyme that can synthesise a new strand of DNA is called DNA polymerase.
what are the 2 types of DNA synthesis
the 2 basic types of DNA synthesis are:
- semi-discontinuous replication
- repair synthesis
what does DNA polymerase do, step by step
DNA polymerase is the enzyme that is responsible for the DNA synthesis of both the leading and lagging strands of the DNA molecule.
DNA polymerase acts in 4 steps:
1.Incoming Nucleotide
2.Nucleophilic attack on alpha-phosphate
3.Formation of phosphodiester bond
4.Pyrophosphate formed as by-product
what are key features of DNA polymerase
- They always need a template
- DNA polymerase catalyses the formation of the covalent bond between 2 nucleotides in the 5’ to 3’ direction.
- DNA polymerases can only add nucleotides to the 3’ end of an existing DNA strand. (They use the free -OH group found at the 3’ end as a “hook,”)
- DNA polymerase is not able to initiate this reaction on its own thus they need an RNA primer with a free 3’OH already base-paired to the template or an existing DNA strand to synthesise the DNA.
- They proofread, or check their work, removing the vast majority of “wrong” nucleotides that are accidentally added to the chain
what are the 5 different DNA polymerases in E.coli and what are they used for
Prokaryotic DNA polymerases:
1. DNA Polymerase I: major repair enzyme
2. DNA Polymerase II: replication restart
3. DNA Polymerase III: principal DNA replication enzyme (Replicase)
4. DNA Polymerase IV: functions in repair
5. DNA Polymerase V: functions in repair
DNA polymerase I and III are involved in DNA replication.
DNA polymerase II, IV and V are involved in DNA repair.
how many DNA polymerases are there in eukaryotes and where are they used
In human cells, there are at least 12 polymerases or more.
- DNA polymerases alpha, delta and epsilon are the main polymerase involved in DNA replication in the nucleus.
- DNA polymerase beta is involved in DNA repair.
- DNA polymerase gamma is the polymerase involved in the synthesis of DNA in the mitochondria.
which DNA polymerase was first discovered
The first DNA polymerase that was discovered was DNA polymerase I.
what 2 parts can a DNA polymerase be cleaved into
DNA polymerase I is a single polypeptide that can be cleaved into 2 parts:
- a large cleaved product, which is also known as a Klenov fragment
- a small cleaved product
which direction does the small and large cleaved products exonuclease activity occur
The small cleaved product has polymerase activity in the 5’ > 3’ direction and exonuclease activity in the 5’ > 3’ direction.
the large cleaved product/Klenov fragment exonuclease activity occurs in the 3’ > 5’ direction.
what is the function of the 5’-3’ exonuclease activity
The 5’ -3’ exonuclease activity is used to remove RNA primers in a reaction called “nick translation.”
To initiate DNA replication, there needs to be attached a nucleotide to the free 3-OH which is present in RNA primer. The RNA primer is made up of another enzyme called RNA primase. At the end of DNA replication, the exonuclease activity of the DNA polymerase removes the short RNA primers on both lagging and leading strands.
