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DNA topoisomerase or simply Topoisomerase (EC 22.214.171.124, type I DNA topoisomerase, untwisting enzyme, relaxing enzyme, nicking-closing enzyme, swivelase, omega-protein, deoxyribonucleate topoisomerase, DNA gyrase) is an enzyme with systematic name DNA topoisomerase. This enzyme catalyzes the following chemical reaction
- Forming a transient phosphodiester bond between a tyrosine residue in the enzyme and one of the ends of a broken strand of DNA.
The overall function of DNA topoisomerase is to manage the topological state of the DNA in the cell. There are two types or families of this enzyme; type I family and type II family. Type I family passes one strand of the DNA through a break in the opposing strand. In other words, DNA topoisomerase type I enzyme cleaves only one strand of DNA. Type II family passes a region of duplex (two strands) from the same molecule or a different molecule through a double stranded gap. To summarize, type II cleaves both strands of DNA, that results in a double-stranded break. Topoisomerases can either relieve negative supercoils, both positive and negative supercoils, or induce positive and negative supercoiling in DNA. The enzymes also can promote catenation (when two single circular DNA strands are linked together after replication) and decatenation (the separation of two linked, closed, circular chromosomes), and can also relieve entanglement of linear chromosomes.
Topoisomerases can be further classified into subfamilies. In the type I family, there are two subfamilies; type IA and type IB when the enzyme links to the 5’ phosphate of the DNA strand, and the 3’ phosphate on the DNA, respectively. In the type II family, the structure and organism determine the subfamilies and their functions.
DNA topology is the tertiary conformations of DNA, such as supercoiling, knotting, and catenation. Topology of DNA can be disrupted by most metabolic processes: RNA polymerase can cause positive supercoils by over-winding the DNA in front of the enzyme, and can also cause negative supercoils by under-winding the DNA behind the enzyme. DNA polymerase has the same effect in DNA replication. Positive and negative supercoiling balance out the entire global topology of the DNA, so overall, the topology remains the same. However, as the DNA replication or transcription fork moves forward and positive supercoiling increases, the DNA strands wrap tighter and tighter around each other, making it more difficult for the polymerase to move forward. It is important for the local topology of DNA ahead of and behind the polymerase to be relieved so that replication and cell division can proceed. This is what DNA topoisomerases are used for.
DNA Topoisomerase Type I Family
DNA Topoisomerase Type I family consists of two subfamilies; type IA and type IB. Type IA DNA topoisomerase amongst various organisms generally share the following properties: All of the enzymes are monomers. The enzyme shares a covalent interaction of a 5' phosphodiester bond at its tyrosine active site with the end of a DNA strand. The mechanism of relaxation of supercoiling requires magnesium(II). In plasmid DNA, the negative supercoils produced can be substrates for the relaxation mechanism, a process which does not go to completion. Type IA also requires an exposed single-stranded region within the DNA substrate. The linking number of DNA changes with relaxation. And type IA topoisomerase can catalyze catenation, decatenation, knotting and unknotting of the DNA.
There are three classes within the subfamily of type IB topoisomerase: topoisomerase I in eukaryotes, topoisomerase V in prokaryotes, and the poxvirus topoisomerase. Type IB subfamily topoisomerases are generally classified by their ability to relax both negative and positive supercoils, and the relaxation mechanism goes to completion (unlike type IA). They form a covalent interaction through the tyrosine active site on the enzyme and the 3’ phosphate on the DNA strand. The relaxation mechanism does not require magnesium(II). Both type IA and type IB topoisomerases, within the Type I family, have very distinct differences in their properties.
DNA Topoisomerase Type II Family
Type II family of topoisomerases share general features and properties that make them distinguishable from the type I family. All type II DNA topoisomerases are dimers. They bind to a duplex DNA and cleave both strands, staggering four bases. Cleavage is done by a covalent interaction between each dimer subunit to the 5’ phosphate on the DNA, creating a phosphotyrosine bond. The reaction pulls the two ends of the cleaved DNA apart – this is called the gated (G-) segment. The transported (T-) segment, a region on the same or a different DNA duplex, is passed through G-segment. This changes the linking number when the DNA is circular (plasmid). The relaxation mechanism requires magnesium(II) and hydrolysis of ATP. The active site containing amino acid tyrosines have a helix-turn-helix motif, which collaborates with acidic residues to activate catalysis. Prokaryotic type II topoisomerases are heterotetrameric. Eukaryotic type II topoisomerases are homodimeric. Type II are also better at DNA relaxation, instead of decatenation, meaning they are better enzymes for relieving topological stresses in linear DNA versus circular DNA.
The types and functions of DNA topoisomerases vary depending on the species and cell type. Yeast, higher eukaryotes, eubacteria, and archaebacteria use different topoisomerase enzymes in DNA replication and transcription.
Yeast cells are known to use three topoisomerases: Topoisomerase I, from the IB subfamily, is required for growth. It provides the replication fork with the ability to move forward, as well as removes positive and negative supercoils associated with transcription. Topoisomerase II from the IIA subfamily, is needed for decatenation of linked chromosomes and preparation for segregation during mitosis. Topoisomerase II cannot induce negative supercoils, but can relax both positive and negative supercoils like topoisomerase I, and can replace topoisomerase I if absent. Topoisomerase III from the IA family is used for cell growth. Without topoisomerase III, recombination rates in mitosis and meiosis can increase, which slows growth in cells. In S. pombe cells, III is used to sustain cell division.
Higher eukaryotic organisms are more complex organisms and typically require more complex cellular machinery. These organisms generally have a topoisomerase I, two type IIA topoisomerases, and two type III enzymes. Topoisomerase I helps with replication fork movement and relaxes supercoils associated with transcription. It is also used for relaxing solenoidal supercoils that form when chromosomes condense in preparation for mitosis. The two type IIA topoisomerases, IIα and IIβ, are used to unlink intertwined daughter duplexes, as well as assist in cell division and suppression of recombination, respectively. Type IIIα and IIIβ are thought to work in embryogenesis and interact with helicases, respectively.
E. coli contains four DNA topoisomerases: two type IA enzymes (I and III), and two type IIA (DNA gyrase and topoisomerase IV). DNA topoisomerase III and IV have similar functions. Topoisomerase III is incapable of relaxing positive supercoils, but it works to support replication fork movement on plasmid DNA in vitro. It can decatenate the winding that is happening behind the replication fork by focusing on nicks in the DNA. Topoisomerase IV is the most effective decatenating enzyme in E. coli. It also relaxes negative supercoils. DNA gyrase uses the hydrolysis of ATP to generate negative supercoiling in bacterial chromosomes. It relaxes positive supercoils ahead of the replication fork and acts in chromosome condensing. Finally, topoisomerase I helps with generating some negative supercoiling along with topoisomerase IV and DNA gyrase.
There is limited knowledge about the archaebacterial genome sequences. Therefore, there is also limited knowledge about the topoisomerase enzymes. They do contain a reverse gyrase, a type IA topoisomerase, and topoisomerase VI. The functions of the topoisomerases in archaebacteria are comparable to the enzymes in eubacteria. The only noteworthy difference is that topoisomerase VI in archaebacteria is responsible for decatenation of DNA replication intermediates, and it relaxes both positive and negative supercoils.
- Gellert M (1981). "DNA topoisomerases". Annual Review of Biochemistry. 50: 879–910. doi:10.1146/annurev.bi.50.070181.004311. PMID 6267993.
- Champoux, James J. (2001). "DNA TOPOISOMERASES: Structure, Function, and Mechanism". Annual Review of Biochemistry. 70: 369–413. doi:10.1146/annurev.biochem.70.1.369. PMID 11395412.CS1 maint: uses authors parameter (link)
- Neuman, Keir C., Seol, Yeonee (November 2016). "The dynamic interplay between DNA topoisomerases and DNA topology". Biophysical Reviews. 8 (Suppl 1): 101–111. doi:10.1007/s12551-016-0240-8. PMC 5418509. PMID 28510219.CS1 maint: uses authors parameter (link)
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