So just a quick recap –
Fundamental rules for DNA replication:-
- DNA replication is semiconservative
- Starts from an origin and proceeds bidirectionally
- Takes place in 5’ to 3’ direction
- Semi-discontinuous process
- Highly accurate
DNA pol III holoenzyme
- Complex of 10 proteins including 2 core enzymes each made up of 3 protein subunits –
- α – polymerisation and proof reading activity
- ε – 3’ to 5’ exonuclease activity
- Θ – stimulates the above
- Another subunit called tau (τ) links the 2 core enzymes. It also activates DnaB helicase enzyme
- Associated with the 2 core enzymes is a subunit – β sliding clamp that allows sliding of the holoenzyme on the DNA. It can only be inserted through nicked DNA molecule where it can recognize 3’-OH ends. It tethers the core enzyme to the DNA strands
- Another complex of proteins is the γ complex or the clamp loader that loads the β sliding clamp on the DNA. It has 5 subunits –
- δ’ – stimulates clamp loading
- δ – function unknown
- γ – binds ATP
- ζ – interacts with SSBP to allow the removal of primase from primers
- ψ – function unknown
DNA pol I holoenzyme
- Constitutes 928 amino acids (AA) produced by pol A gene
- Has 3 major activities
- 5’ to 3’ exonuclease activity
- 5’ to 3’ polymerase activity
- 3’ to 5’ exonuclease activity
- Has 2 major domains that can be broken down by proteolysis by trypsin or subtilin
- Smaller N-terminal has 5’ to 3’ exonuclease activity
- Larger C-terminal has 5’ to 3’ polymerase and 3’ to 5’ exonuclease activity. This is known as the Klenow fragment
DNA pol IV & V
- They induce polymerization in case of DNA damage. Hence, they are responsible for 50% mutations
- Responsible for low fidelity of DNA replication as they confer the potential to survive with considerable amount of mutations to the DNA
- Have low processivity
- Pol IV is encoded by dinB gene
- Pol V is encoded by umu C/D gene
Compare & Contrast – DNA pol I and III
Compare:
- Both are involved in chromosome replication process
- Both are highly accurate
- Have proof-reading activity
- Have 5’ to 3’ polymerase and 3’ to 5’ exonuclease activity
Contrast:
|
DNA Pol I |
DNA Pol III |
| Involved in chromosome replication and DNA repair |
Involved only in chromosome replication |
|
Removes primer after DNA synthesis is completed and fills in the gaps |
Polymerizes new DNA strands by adding nucleotides |
| It also has 5’ to 3’ exonuclease activity apart from 5’ to 3’ polymerase and 3’ to 5’ exonuclease activities |
Has only 5’ to 3’ polymerase and 3’ to 5’ exonuclease activities |
|
Made up of a single polypeptide |
It has a core enzyme made up of 3 polypeptides with 6 additional polypeptides |
|
Not very processive |
Highly processive |
|
Adds 20-100 nucleotide per event |
Adds 850 nucleotides per event |
|
There are about 10-20 molecules in a cell |
There are about 400 molecules of this enzyme in a cell |
|
Encoded by pol A gene |
Encoded by pol C gene |
|
Characteristics |
DNA pol I | DNA pol II | DNA pol III |
| Encoded by | Pol A gene | Pol B gene |
Pol C gene |
|
No.of subunits |
1 | >4 | >10 |
|
3’ to 5’ exonuclease activity |
yes | yes | yes |
| 5’ to 3’ exonuclease activity | Yes | no |
no |
|
Polymerase rate |
16-20 nucleotides/second | 5-10 nucleotides/second | 250-1000 nucleotides/second |
| Processivity | low | low |
high |