It also requires a free 3'-OH group to which it can add nucleotides by forming a phosphodiester bond between the 3'-OH end and the 5' phosphate of the next nucleotide. Topoisomerase and SSB proteins are important components of the replication process in prokaryotes, and there are similar proteins that are also found in eukaryotes. As the DNA-binding gate separates, the T-segment is transferred through the G-segment. Unlike type IIA topoisomerases, type IIB topoisomerases cannot simplify DNA topology (see below), but they share several structural features with type IIA topoisomerases. The first type II topoisomerase to be discovered was DNA gyrase from bacteria, by Martin Gellert and coworkers in 1976,[5] and also characterized by Nicholas Cozzarelli and co-workers. These signal-regulated genes include genes activated in response to stimulation with estrogen, serum, insulin, glucocorticoids (such as dexamethasone) and activation of neurons. [52][53][54][55] Topo II, with other associated enzymes,[54] appears to be important for the release of paused RNA polymerase at highly transcribed or long genes. the transcription start site of a gene. often found in intergenic regions. The nucleosome was close to the transcription start site of the gene. In addition, drug-resistant bacteria often have a point mutation in gyrase (Serine79Alanine in E. coli) that renders quinolones ineffective. [56][57][58] For the genes at which it occurs, the DNA double-stranded break induced by TOP2B is thought to be part of the process of regulation of gene expression. Studies have suggested that this region is regulated by phosphorylation and this modulates topoisomerase activity, however more research needs to be done to investigate this. The structure of topo VI-A was solved by Bergerat et al. [55] The components of the non-homologous end joining DNA repair pathway were essential to the closing of the DNA double-strand break.[52]. It is noteworthy that type IB topoisomerases are found in some bacteria . DNA topoisomerases: structure, function, and mechanism The second gene, termed topo VI-A (Pfam PF04406), contains the WHD and the Toprim domain. 5). Please expand the section to include this information. The G segment is part of a much longer piece of DNA (>100 bp) that is wrapped around the enzyme, one arm of which forms the T-segment that is passed through the double-stranded break (Fig. 10.4: The Structure and Function of Cellular Genomes Skills to Develop Explain the process of DNA replication in prokaryotes Discuss the role of different enzymes and proteins in supporting this process DNA replication has been extremely well studied in prokaryotes primarily because of the small size of the genome and the mutants that are available. Type IC topoisomerases form a covalent 3'-phosphotyrosine intermediate. [4] Biochemistry, electron microscopy, and recent structures of topoisomerase II bound to DNA reveal that type IIA topoisomerases bind at the apices of DNA, supporting this model. The other strand, complementary to the 5' to 3' parental DNA, is extended away from the replication fork, in small fragments known as Okazaki fragments, each requiring a primer to start the synthesis. There are a number of protein inhibitors of gyrase, including the bacterial toxins CcdB, MccB17, and ParE,[38][39][40] that stabilize the cleavage complex, in a similar manner to FQs. The break is then separated, using domain II as a hinge, and a second duplex or strand of DNA is passed through. The first report of the presence of a type IB DNA topoisomerase in prokaryotes. In these complexes, topoisomerases I and II create single- and double-stranded breaks, respectively. This is a mathematical identity originally obtained by Clugreanu in 1959[16] and is referred to as the Clugreanu, or ClugreanuWhiteFuller, theorem. . The enzyme first wraps around DNA and creates a single, 3' phosphotyrosine intermediate. This assemblage was all present at the linker DNA adjacent to a single nucleosome in the promoter region of a gene (see Figure). Type IB topoisomerases were originally identified in eukaryotes and in viruses. Topoisomerases are ubiquitous enzymes, found in all living organisms.[1]. They use the hydrolysis of ATP, unlike Type I topoisomerase. The G-segment is sealed, leading to the C-terminal gate (or C-gate) to open, allowing for the release of the T-segment. Type IIB topoisomerases operate through a similar fashion, except that the protein forms a two-base overhang in the G-segment and the C-terminal gate is completely missing. Solved Type I topoisomerases O pass one strand of a DNA - Chegg Metrics Abstract Human topoisomerases comprise a family of six enzymes: two type IB (TOP1 and mitochondrial TOP1 (TOP1MT), two type IIA (TOP2A and TOP2B) and two type IA (TOP3A and TOP3B). This last structure showed that the Toprim domain and the WHD formed a cleavage complex very similar to that of the type IA topoisomerases and indicated how DNA-binding and cleavage could be uncoupled, and the structure showed that DNA was bent by ~150 degrees through an invariant isoleucine (in topoisomerase II it is I833 and in gyrase it is I172). Type IA topoisomerases operate through a strand-passage mechanism, using a single gate (in contrast with type II topoisomerases). Drugging Topoisomerases: Lessons and Challenges - PMC supercoiling is an energy-requiring process. They do this by binding to DNA and cutting the sugar-phosphate backbone of either one (type I topoisomerases) or both (type II topoisomerases) of the DNA strands. Helicase opens up the DNA-forming replication forks; these are extended bidirectionally. It turns out that there are specific nucleotide sequences called origins of replication where replication begins. This strand passage mechanism shares several features with type IIA topoisomerases. These structures, of which one is an X-ray crystal structure and the other is a Small-Angle X-ray Scattering (SAXS) reconstruction, show that the ATPase domain can be either open or closed. It turns out that there are specific nucleotide sequences called origins of replication where replication begins. Involved in DNA repair. In the strand passage mechanism, the cleavage of DNA is key to allow the T-segment to transfer through the G-segment. [50][51], At least one topoisomerase, DNA topoisomerase II beta (topo II), has a regulatory role in gene transcription. The strand-passage reaction can be intra- or intermolecular (Fig. Type II topoisomerases are found in organisms of all domains of life and are encoded in most, except for a few extremely reduced ones, . Originally found in archaea, they have also been found in eukaryotes, and, in particular, in plants; examples include topo VI and topo VIII. Along with gyrase, most prokaryotes also contain a second type IIA topoisomerase, termed topoisomerase IV. DNA topoisomerases: Advances in understanding of cellular roles and This is a highly-effective mechanism of inhibition that is also used by several topoisomerase-targeted anti-cancer drugs. [27] This controlled-rotation mechanism was first described for Vaccinia topo I[27][28] and permits DNA rotation of the free end around the intact strand, the speed being controlled by 'friction' within the enzyme cavity, before the nick is re-ligated (Fig. [12] Gp55.2 binds DNA and specifically blocks the relaxation of negatively supercoiled DNA by topoisomerase I. The catalytic tyrosine lies on this WHD. Prevents excessive supercoiling of the genome, and supports transcription, Removes (-), but not (+) supercoils; overlapping function with topoisomerase IV, Removes (-), but not (+) supercoils; assists in the unlinking of precatenanes in cellular DNA replication; can catalyze the knotting, unknotting, and interlinking of single-stranded circles as well as the knotting, unknotting, catenation, and decatenation of gapped or nicked duplex DNA circles, Has been shown to be a putative RNA topoisomerase. Type IA topoisomerases form a covalent linkage between the catalytic tyrosine and the 5'-phosphoryl. Type IC topoisomerases share a similar mechanism to the type IB enzymes but are structurally distinct. In contrast to type I topoisomerases that are generally monomeric, type II topoisomerases are homodimers or heterotetramers. DNA topology refers to the crossing of the two DNA strands that alters the twist of the double helix and gives rise to tertiary conformations of DNA, such as supercoils, knots and catenanes. During a DNA relaxation reaction this process changes the linking number of the DNA by +/-1 (Fig. Topoisomerase 1 inhibition is synthetically lethal with deficiency of expression of certain DNA repair genes. [30] Further, in the absence of ATP, gyrase is able to remove negative supercoils in a slower DNA relaxation reaction. Type 1 topoisomerase is inhibited by irinotecan, topotecan, hexylresorcinol and camptothecin. It was first found by J.C. Wang in the 1970s while working on Escherichia coli. In this process, these enzymes change the linking number of circular DNA by 2. A recent structure of the topo VI A/B complex was solved, showing an open and closed conformation, two states that are predicted in the two-gate mechanism (see below). [56][57][58], Stimulus-induced DNA double-strand breaks (DSBs) that are limited to a short-term (10 minutes to 2 hours) are induced by topo II in the promoter regions of signal-regulated genes. They introduce a transient single-stranded break through the formation of a tyrosyl-phosphate bond between a tyrosine in the enzyme and a 5-phosphate in the DNA. A protein called the sliding clamp holds the DNA polymerase in place as it continues to add nucleotides. they prevent completion of the catalytic cycle of topo II but do not stabilize the DNA cleavage complex. The majority of topo-targeted drugs act in this way, i.e. Bacteriophage T4 gene 39", "The DNA-delay mutants of bacteriophage T4", "The genetics of the Luria-Latarjet effect in bacteriophage T4: evidence for the involvement of multiple DNA repair pathways", "Topoisomerase involvement in multiplicity reactivation of phage T4", "Formation of covalently closed heteroduplex DNA by the combined action of gyrase and RecA protein", https://en.wikipedia.org/w/index.php?title=Type_II_topoisomerase&oldid=1139880087, CS1 maint: DOI inactive as of December 2022, Articles to be expanded from October 2021, Articles with unsourced statements from November 2014, Creative Commons Attribution-ShareAlike License 4.0, Type IIA topoisomerases include the enzymes. [49], Etoposide (Fig. These compounds work by interacting with their target (gyrase or topo IV) and DNA at the cleavage site to stabilize the DNA-protein covalent cleavage intermediate. Topo IIdependent double-strand DNA breaks and components of the DNA damage repair machinery are important for rapid expression of immediate early genes, as well as for signal-responsive gene regulation. They change the topology of DNA via transient breaks on either one or two of the DNA strands to allow passage of another single or double DNA strand through the break. [10] They do this via transient breakage of one or both strands of DNA. Topoisomerases as Anticancer Targets - PMC - National Center for This page was last edited on 23 September 2021, at 01:24. 4). The consequences of topological perturbations in DNA are exemplified by DNA replication during which the strands of the duplex are separated; this separation leads to the formation of positive supercoils (DNA overwinding or overtwisting) ahead of the replication fork and intertwining of the daughter strands (precatenanes) behind[10][19] (Fig. Topoisomerase II: Cleaves both DNA strands for larger structural alterations of DNA; Requires ATP 14.4.1: DNA Replication in Prokaryotes - Biology LibreTexts [6] DNA gyrase catalyzes the introduction of negative supercoils into DNA and is the only type II enzyme to do this, all the others catalyze DNA relaxation. Molecular mechanism of DNA replication (article) | Khan Academy You isolate a cell strain in which the joining together of Okazaki fragments is impaired and suspect that a mutation has occurred in an enzyme found at the replication fork. Topoisomerase: Overview & Applications - Excedr How does the replication machinery know where to begin? Domain III and I close and the DNA is re-annealed. The 3 parameters are related as follows: Lk = Tw +Wr. Enzyme turnover requires the binding and hydrolysis of ATP. The process of DNA replication can be summarized as follows. As pointed out by Singh et al.,[58] "about 80% of highly expressed genes in HeLa cells are paused". [2] Indeed, these enzymes are of interest for a wide range of effects. The process is quite rapid and occurs without many mistakes. Following traversal . 1. This was eventually substantiated by the Dong et al. They are classified into two subtypes based on evolutionary, structural, and mechanistic considerations. amino terminal 9kda domain of vaccinia virus dna topoisomerase i residues 1-77, experimental electron density for residues 1-77, Please expand the article to include this information. Linear DNA in eukaryotes is so long they can be thought of as being without ends; type II topoisomerases are needed for the same reason. The . PMID: 25875362; PMCID: PMC4396842, "Crystal structure of the amino-terminal fragment of vaccinia virus DNA topoisomerase I at 1.6 A resolution", "Structure of the N-terminal fragment of topoisomerase V reveals a new family of topoisomerases", "Topoisomerase V relaxes supercoiled DNA by a constrained swiveling mechanism", "Drugging topoisomerases: lessons and challenges", "Epigenetic inactivation of the premature aging Werner syndrome gene in human cancer", "MRE11-deficiency associated with improved long-term disease free survival and overall survival in a subset of stage III colon cancer patients in randomized CALGB 89803 trial", "ATM expression predicts Veliparib and Irinotecan sensitivity in gastric cancer by mediating P53 independent regulation of cell cycle and apoptosis", "Identification of cetrimonium bromide and irinotecan as compounds with synthetic lethality against NDRG1 deficient prostate cancer cells", "mTOR regulates the expression of DNA damage response enzymes in long-lived Snell dwarf, GHRKO, and PAPPA-KO mice", https://en.wikipedia.org/w/index.php?title=Type_I_topoisomerase&oldid=1045901823, Articles to be expanded from February 2021, Creative Commons Attribution-ShareAlike License 4.0. Their catalytic intermediates, the . DNA replication and repair - Knowledge @ AMBOSS Type IIB enzymes show important structural differences, but are evolutionarily related to the type IIA enzymes. RNA polymerase II frequently has a pausing site that is about 3060 nucleotides downstream of Relaxation is not an active process and energy (in the form of ATP) is not spent during the nicking or ligation steps; this is because the reaction between the tyrosine residue at the active site of the enzyme with the phosphodiester DNA backbone simply replaces one phosphomonoester bond with another. structure that was solved in the presence of DNA. This domain is thought to communicate the nucleotide state of the ATPase domain to the rest of the protein. [17][18] Lk cannot be altered without breaking one or both strands of the helix; Tw and Wr are interconvertible and depend upon the solution conditions. Involved in RNA processing, Removes (-), but not (+) supercoils, introduces positive supercoils, Removes (+) and (-) supercoils; supports fork movement during replication and transcription. DNA polymerase can only extend in the 5' to 3' direction, which poses a slight problem at the replication fork. [16] The nucleoprotein complex was captured with a long DNA duplex and gepotidacin, a novel bacterial topoisomerase inhibitor. Reverse gyrase is particularly interesting because an ATPase domain, which resembles the helicase-like domain of the Rho transcription factor, is attached (the structure of reverse gyrase was solved by Rodriguez and Stock, EMBO J 2002). DNA topoisomerases (or topoisomerases) are enzymes that catalyze changes in the topological state of DNA, interconverting relaxed and supercoiled forms, linked (catenated) and unlinked species, and knotted and unknotted DNA. [4], DNA topoisomerases are divided into two classes: type I enzymes (EC; topoisomerases I, III and V) break single-strand DNA, and type II enzymes (EC; topoisomerases II, IV and VI) break double-strand DNA.[5]. Step by step Solved in 2 steps See solution Check out a sample Q&A here Knowledge Booster Learn more about The structures of the N-terminal ATPase domain of gyrase[8] and yeast topoisomerase II[9] have been solved in complex with AMPPNP (an ATP analogue), showing that two ATPase domains dimerize to form a closed conformation. These enzymes catalyze changes in DNA topology via transient single-stranded breaks in DNA. DNA topoisomerases in cancer therapy - PubMed Type IIA topoisomerases have the special ability to relax DNA to a state below that of thermodynamic equilibrium, a feature unlike type IA, IB, and IIB topoisomerases. 14.4: DNA Replication in Prokaryotes - Biology LibreTexts Examples of type IA topoisomerases include prokaryotic topo I and III, eukaryotic topo III and III and the archaeal enzyme reverse gyrase. Function Type II topoisomerases increase or decrease the linking number of a DNA loop by 2 units, and it promotes chromosome disentanglement. [15] The Fass structure shows that the Toprim domain is flexible and that this flexibility can allow the Toprim domain to coordinate with the WHD to form a competent cleavage complex. DNA gyrase - Wikipedia Rather than utilizing a strand-passage mechanism, these enzymes operate via a 'swivel' or 'controlled rotation' of the cleaved strand around the intact strand. Single-strand binding proteins coat the single strands of DNA near the replication fork to prevent the single-stranded DNA from winding back into a double helix. 7) and its close relative teniposide (VM-26) are epipodophyllotoxin derivatives obtained from the rhizome of wild mandrake that target topo II by stabilizing the covalent cleavage complex and preventing religation of the cleaved DNA. [32][33][44][45][46][47] Most of these compounds act in a similar way to FQs, i.e. DNA Topoisomerase - an overview | ScienceDirect Topics 3). 5). Topoisomerases bind to DNA in a noncovalent fashion followed by the formation of transient cleavage complexes. The Toprim fold is a Rossmann fold that contains three invariant acidic residues that coordinate magnesium ions involved in DNA cleavage and DNA religation. All topoisomerases form a phosphotyrosine intermediate between the catalytic tyrosine of the enzyme and the scissile phosphoryl of the DNA backbone. The deficiencies can arise through mutation, epigenetic alteration or by inhibition of a gene's expression. a central DNA-binding core (which structurally forms a heart-shaped structure), and, Inhibitors of type II topoisomerase include, Poisons of type II topoisomerases include, This page was last edited on 17 February 2023, at 09:42. DNA ligase, as this enzyme joins together Okazaki fragments. This results in a variable change of linking number per cleavage and religation event. In E. coli, type I topoisomerase can only relieve negatively supercoiled DNA (negative supercoiling is the end result of newly replicated DNA genome). Both Pfam signatures are found in the single-chain eukayotic topoisomerase. [citation needed], The bacteriophage (phage) T4 gyrase (type II topoismerase) is a multisubunit protein consisting of the products of genes 39, 52 and probably 60. [33] Camptothecin and its derivatives act by stabilizing the topo I cleavage complex, preventing religation of the protein-mediated nick in the DNA. Left unresolved, links between replicated DNA will impede cell division. The general strand-passage mechanism for the type II topos begins with the binding of one DNA duplex, termed the gate segment (G-segment), at the DNA gate. Structure of the 42 KDa fragment of the N-terminal ATPase and transducer domains of DNA gyrase homologous to all other type IIA topoisomerases. The capping lobe and catalytic lobe wrap around the DNA. Type IIA topoisomerases catalyze transient double-stranded breaks in DNA through the formation of tyrosyl-phosphate bonds between tyrosines in the enzyme (one on each subunit) and 5-phosphates staggered by 4 bases in opposite DNA strands. Foreword. Human DNA-Topoisomerases - Diagnostic and Therapeutic - PubMed Topoisomerases are ubiquitous proteins found in all three domains of life. 2). Further details may exist on the, Mattenberger Y, Silva F, Belin D. 55.2, a phage T4 ORFan gene, encodes an inhibitor of Escherichia coli topoisomerase I and increases phage fitness. The two classes of topoisomerases possess a similar strand passage mechanism and domain structure (see below), however they also have several important differences. If left unchanged, this torsion would eventually stop the DNA or RNA polymerases involved in these processes from continuing along the DNA helix. We also acknowledge previous National Science Foundation support under grant numbers 1246120, 1525057, and 1413739. Prokaryotes appear to use their genomes very efficiently, with only an average of 12% of the genome being taken up by noncoding . They are indispensable for the control of DNA topology. An enzyme called helicase unwinds the DNA by breaking the hydrogen bonds between the nitrogenous base pairs. Once replication is completed, the RNA primers are replaced by DNA nucleotides and the DNA is sealed with DNA ligase, which creates phosphodiester bonds between the 3'-OH of one end and the 5' phosphate of the other strand. they stabilize the enzyme-DNA covalent cleavage intermediate.[32][33][34]. When helicase and topoisomerase meet! - PubMed [29] Since the host E. coli DNA gyrase can partially compensate for the loss of the phage T4 gene products, mutants defective in either genes 39, 52 or 60 do not completely abolish phage DNA replication, but rather delay its initiation. [9] Type IC topoisomerases work through a controlled rotary mechanism, much like type IB topoisomerases[10](pdb ID = 2CSB and 2CSD), but the fold is unique. Knots in DNA can be found in bacteriophages and as products of recombination reactions. Recent single molecule experiments have confirmed what bulk-plasmid relaxation experiments have proposed earlier, which is that uncoiling of the DNA is torque-driven and proceeds until religation occurs. These interfacial inhibitors are stabilized by stacking interactions with the nicked DNA and hydrogen bonding to the enzyme. In prokaryotes, gyrase is an antibacterial target. Roles of eukaryotic topoisomerases in transcription - Nature As type II topoisomerses break a double strand, they can fix this state (type I topoisomerases could do this only if there were already a single-strand nick), and the correct chromosome number can remain in daughter cells. V Perspectives DNA topoisomerases have been identified as the cellular targets of many potent antitumor drugs. Topoisomerase binds at the region ahead of the replication fork to prevent supercoiling. This ability, known as topology simplification, was first identified by Rybenkov et al. [10] Potential topological issues associated with the double-helical structure of DNA were recognized soon after its structure was first elucidated in 1953 by James Watson, Francis Crick and Rosalind Franklin[11][12][13] and developed further by the work of Max Delbruck and John Cairns. [14] and one by Dong et al. The sliding clamp is a ring-shaped protein that binds to the DNA and holds the polymerase in place. These include YacG[41] and pentapeptide repeat proteins, such as QnrB1 and MfpA;[42][43] these protein inhibitors also confer resistance to fluoroquinolones. Extrachromosomal DNA in eukaryotes includes the chromosomes found within organelles of prokaryotic origin (mitochondria and chloroplasts) that evolved by . First, the single-stranded DNA binds domain III and I. Full article: Type IA topoisomerases can be "magicians" for both DNA A coiled-coil region leads to a C-terminal domain that forms the main dimer interface for this crystal state (often termed the C-gate). Type IA topoisomerases, historically said to be found in prokaryotes, create a single break in DNA and pass a second strand or duplex through the break. Although these proteins are not viable as antibacterials, their mode of action could inspire the development of novel antibacterial compounds. [55] Topo II and PARP-1 increased at the site of the double-strand break and components of the non-homologous end joining DNA repair pathway, including DNA-PKcs, Ku70/Ku80 and DNA ligase IV assembled with topo II and PARP-1. In prokaryotes, gyrase is an antibacterial target. The human topoisomerase type IB enzyme forms a covalent 3'-phosphotyrosine intermediate, the topoisomerase 1-cleavage-complex (Top1cc). The first structure of a C-terminal domain of gyrase was solved by Corbett et al. [19] showing that the HTH and Toprim fold had a novel conformation compared with that of topo IIA. Last updated on September 19, 2022 by Excedr Topoisomerase: Overview Topoisomerase (DNA topoisomerases) is an enzyme that catalyzes the changes in the intertwined state of two DNA strands. Eukaryotic Top 1 belongs to the type IB subtype, and topoisomerase V found in the archaeal genus Methanopyrus is, thus far, the only member of the type IC subtype . The gap between the two DNA fragments is sealed by DNA ligase, which helps in the formation of phosphodiester bonds. The topoisomerase II core was later solved in new conformations, including one by Fass et al. The enzyme uses the hydrolysis of ATP to introduce positive supercoils and overwinds DNA, a feature attractive in hyperthermophiles, in which reverse gyrase is known to exist.