Introduction and application of DNA polymerase properties

DNA polymerase, also known as DNA-dependent DNA polymerase (DNA pol), is a class of enzymes
that use parent DNA as a template to catalyze the polymerization of substrate dNTP molecules to form offspring DNA 。 This enzyme was first discovered by American scientist Arthur Komberg in Escherichia coli in 1957, known as DNA polymerase I.
(DNA polymerase I.
, referred to as polI.
) and has been found in other prokaryotes and eukaryotes
。 The common features of these DNA polymerases are: (1) have 5' →3' polymerase activity, which determines that DNA can only be synthesized in the 5' →3' direction; (2) Primers are required, and DNA polymerase cannot catalyze the synthesis of new DNA strands from scratch, but can only catalyze dNTP to join the 3'-OH end
of the nucleotide chain.
Therefore, the replication initially requires a 3'-OH end of the DNA primer as the starting point to synthesize a new strand
in the 5' →3' direction.
First, the characteristics:
THERE are many kinds of DNA polymerases, E.
coli has three kinds
.
Usually DNA polymerases have the following common characteristics:
1, the need for DNA templates, so this type of enzyme is also called DNA-dependent DNA polymerase;
2, the need for RNA or DNA as primer (primer), that is, DNA polymerase can not be catalyzed from scratch;
3, catalytic dNTP added to the 3'-OH end of the primer, its rate is 1000nt/min, so the direction of DNA synthesis is 5' to 3';
4, the three DNA polymerases are multifunctional enzymes, they play a role
in different stages of the DNA replication and repair process.
Application:
E.
coli's DNA pol I.
involves DNA damage repair and plays an auxiliary role
in semi-reserved replication.
DNA polII also plays an important role
in repairing damage.
DNA pol III.
is a multi-subunit protein that acts as a replicase in the de novo synthesis of new strands
of DNA.
The problem of replication fidelity affects the accuracy of translation, which relies heavily on the specific pairing of bases
.
It is estimated that each base pair will have ^10-3
.
There is a probability of mismatch, but the actual mismatch rate is only ^10-8 to ^10-10, that is, only one error per genome in every 1000 bacterial replication cycles
.
DNA polymerase can increase the specificity of complementary bases, which is mainly manifested in the following two aspects: 1, can check whether the incoming base is complementary to the template, then identified by a matching chemical characteristic, which is a preventive measure before synthesis, called presynthetic error control;
2.
After the new base is added to the chain, check whether the bases are paired
.
If a mismatch occurs, the wrong base just added will be removed, which is proofreading
.
The bacteria's three DNA polymerases all have 3 7 nitro excision activity, which is processed against the direction of DNA synthesis, providing proofreading function
.
In the extension phase of the chain, a nucleotide enters the end of the human growth chain, forming a bond, and the enzyme moves forward one base pair to prepare for the entry
of the next base pair.
If an error occurs, the enzyme retreats, excising the base that was added last, creating a site that is then replaced
by the correct base.
The relationship between polymerization and correction of different DNA polymerases is different
.
Sometimes these activities are produced by the same protein subunits, but sometimes they have different subunits
.
The elimination of an erroneous base is actually quite complex because the excision reaction is catalyzed by different sites
.