DNA polymerase, also known as DNA dependent DNA polymerase (DNA pol), takes parental DNA as template and catalyzes substratedNTPA class of enzymes that polymerize molecules to form progeny DNA.This enzyme was originallyU.S.AscientistArthur Komberg in 1957Escherichia coliDNA polymerase I (pol I for short) has been found in other prokaryotes and eukaryotes.The common characteristics of these DNA polymerase are: ① they have 5 '→ 3' polymerase activity, which determines that DNA can only be synthesized along the 5 '→ 3' direction; ②needprimerDNA polymerase can not catalyze the de novo synthesis of new DNA strands, but can only catalyze the addition of dNTP to the 3 '- OH end of the nucleotide chain.Therefore, at the beginning of replication, the 3 '- OH end of an RNA primer is required as the starting point to synthesize a new chain in the direction of 5' → 3 '.[1][7]
In 1953,Watson andCrickHe published a classic paper describing the chemical structure of DNA, but some scientists initially questioned its importance.In their paper, the two proposed that the principle of DNA replication remains to be determined.At that time, American biochemistArthur Kornberg He is working in the Department of Microbiology at Washington University in St. Louis, Missouri, and he recognized the significance of this paper.As a result, he became interested in the process of nucleic acid synthesis, especially DNA synthesis.What he used in these studies is relatively simpleEscherichia coliIn 1956, he discovered the enzyme that assembles the basic unit of DNA.This enzyme is called DNA polymerase I, and it appears in all organisms in several different variants.Koenberg described these findings in his paper, which was rejected at first, but was accepted and published by the famous Journal of Biochemistry in 1957.In 1959, he becameNobel PrizeOne of the co winners of.[2]
The discovery of DNA polymerase I (pol I) is of great significance to biological research, because it plays a central role in the life process, making us realize how DNA replicates and repairs.Before cell division, pol I will replicate all the components of cell DNA.Next, the mother cell passes copies of its DNA to each daughter cell, thus passing on genetic information from generation to generation.Koenberg found that pol I can read the complete DNA strand and use it as a template to synthesize a new strand, which is exactly the same as the original DNA strand - this process is no different from copying documents with a photocopier.[2]
However, the copying machine is mechanical when copying documents, and it does not care about the contents of the documents. In contrast, some members of the seven subclasses of DNA polymerase can check the original DNA template, detect, remove and correct errors, so as to produce an error free new DNA chain, including DNA polymerase I.Other DNA polymerases can only replicate and cannot be repaired, so they can retain mutations in the genome or kill cells.[2]
characteristic
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There are many kinds of DNA polymerase, and E.coli has three kinds.Generally, DNA polymerase has the following common characteristics:[3]
① DNA template is required, so this kind of enzyme is also called DNA dependent DNA polymerase;[3]
② RNA or DNA is required asprimer(prime), that is, DNA polymerase cannot be de novo catalyzed;[3]
③ Catalytic dNTP is added to the 3 '- OH end of the primer at a rate of 1000 nt/min, so the direction of DNA synthesis is 5' to 3 ';[3]
④ All three DNA polymerase belong to multifunctional enzymes, which play roles in different stages of DNA replication and repair process.[3]
type
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prokaryote
Escherichia coli DNA polymerase DNA polymerase was first found in E.coli. So far, five types of DNA polymerase have been identified, namely DNA polymerase I, DNA polymerase II, DNA polymerase III, DNA polymerase IV and DNA polymerase V, which are all related to the extension of DNA strand.[4]
Among them, DNA polymerase I, II and III have been studied clearly.[5]
Comparison of DNA Polymerase Properties in Prokaryotic Cells
DNA Polymerase IIt was first discovered by Arthur Komberg in E.coli in 1956. It is a multifunctional enzyme with three different active centers:[4]
① 5 '- 3' polymerase activity catalyzes the extension of DNA chain, mainly used to fill gaps in DNA or cut offRNA primerThe space left behind;[4]
② The 3 '- 5' exonuclease activity can recognize and cut off the mismatched nucleotides at the 3 'end of DNA during polymerization, and play a role in proofreading;[4]
③ The activity of 5 '- 3' exonase is mainly used to remove 5 'primers or damaged DNA.The mutant with this enzyme defect can still survive, indicating that DNA polymerase I is not the main polymerase for DNA replication.[4]
DNA Polymerase IIIs a kind ofMultienzyme complexThere are 5 '- 3' polymerase active center and 3 '- 5' exonase active center, but there is no 5 '- 3' exonase active center.Its catalytic activity for 5 '- 3' direction synthesis reaction is only 5% of that of DNA polymerase I.Because the DNA replication of the E.coli mutant with the enzyme defect is normal, it is not the main polymerase for DNA replication, and may play a role in DNA damage repair.[4]
DNA Polymerase IIIIt is a multi enzyme complex, and the whole enzyme is composed ofα、β、γ、δ、ε、θ、τ、χandψThere are 10 subunits in total, includingα、εandθThe subunits form the core of the whole enzyme.αThe subunit contains 5 '- 3' polymerase active center,εThe subunit contains 3 '- 5' exonuclease active center,θSubunits may play an assembly role, and other subunits have different roles.DNA polymerase III has the highest activity, plays a leading role in the extension of DNA replication chain, and is the main enzyme that catalyzes DNA replication synthesis.[4]
DNA Polymerase IV and VIt was discovered in 1999 and is mainly involved in DNA repair.[4]
Eukaryote
More than 15 kinds of eukaryotic DNA polymerase have been found.There are mainly five kinds of DNA polymerase in mammalian cells, which are called DNA polymerase respectivelyα、β、γ、δandε，They all have 5 '- 3' polymerase activity.[4]
The DNA polymerase of eukaryotic cells and bacterial DNA polymerase have the same basic propertiesdNTPAs substrate, Mg is required2+Activated. Aggregation must haveTemplate chainAnd the primer chain with 3 '- OH terminal, the extension direction of the chain is 5' → 3 '.However, DNA polymerase of eukaryotic cells generally does not have exonuclease activity, so it is speculated that there must be another enzyme that plays a role in proofreading DNA replication.
DNA PolymeraseαIts main function is primer synthesis, that is, it can start the synthesis of the leading chain and the following chain.It is related toInitiator enzymeThe formation of complex is also called pol because of its dual functions of hair and extension chainαThe initiator enzyme of.DNA PolymeraseβThe activity level is stable, which may mainly play a role in the repair of DNA damage. It belongs to a high fidelity repair enzyme.DNA PolymeraseδIt is an enzyme mainly responsible for DNA replication, participating inLeader chainandTrailing chainSynthesis of.DNA polymeraseεIt is related to post chain synthesis and plays an important role in nucleoside excision and base excision repair in DNA synthesis. Moreover, it may have some functions in cell reorganization.DNA PolymeraseγstayMitochondrial DNAPlay a role in replication.
In addition to the above five major DNA polymerase, there are alsoζ、η、τandκSeveral DNA polymerase are responsible for repairing damage, but their fidelity is very low.[6]
Comparison of DNA Polymerase Properties in Eukaryotic Cells
The fidelity of DNA replication is the guarantee of stable transmission of genetic information.Organisms have at least three mechanisms to achieve fidelity:[4]
① Abide by strict base pairing rules;[4]
② The selection of substrate by the active center of 5 '- 3' polymerase makes the mismatch ratio of nucleotide only 10-4~10-5；[4]
③ Immediate proofreading of 3 '- 5' exonuclease active center in case of replication error, reducing the mismatch rate to 10-6~10-8。[4]
application
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E. Coli's DNA pol I is involved in DNA damage repair and plays an auxiliary role in semi reserved replication.DNA pol Ⅱ also plays an important role in repairing damage.DNA pol Ⅲ is a multi subunit protein, which plays the role of replicase in the de novo synthesis of new DNA strands.[3]
The fidelity of replication will affect the accuracy of translation, which mainly depends on the specific pairing of bases.It is estimated that each base pair will have 10-3。But the actual mismatch rate is only 10-8~10-10That is, in every 1000 bacterial replication cycles, each genome only generates one error.DNA polymeraseIt can increase the specificity of complementary bases, which is mainly shown in the following two aspects:[3]
① It can check whether the incoming base is complementary to the template. At this time, it can be identified by a matching chemical feature, which is a preventive measure before synthesis, called pre synthetic error control;[3]
② After the new base is added to the chain, check whether the base pairs.In case of mismatch, the newly added wrong base will be removed, which is called proofreading.[3]
The three DNA polymerases of bacteria all have 37 nitrate 'exo cutting activity, which is processed against the direction of DNA synthesis to provide proofreading function.In the chain extension stage, a nucleotide enters the end of the long chain to form 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 will go back, remove the last base added, generate a site, and then be replaced by the correct base.[3]
The relationship between polymerization and calibration of different DNA polymerase is different.Sometimes, these activities are caused by the sameProtein subunitBut sometimes they have different subunits.The elimination of a wrong base is actually very complicated, because this excision reaction is catalyzed by different sites.[3]
