Horizontal gene transfer (HGT) between prokaryotic organisms, such as bacteria, is a common way to generate evolutionary innovations. Type IC differs from type IB or any other known topoisomerase in sequence or structure, with a unique fold in its N-terminal domain [47]. government site. Molecular dissection of Helicobacter pylori Topoisomerase I reveals an additional active site in the carboxyl terminus of the enzyme. CRISPR/Cas9-based deletion that targeted the TOP3BTDRD3 complex did not affect Flavivirus translation and replication, but was found to diminish the late-stage production of infectious virus particles [37]. 2023 Jan 9;16(1):94. doi: 10.3390/ph16010094. Nurse P., Levine C., Hassing H., Marians K.J. Bizard A.H., Hickson I.D. This mutation was utilized to obtain the crystal structure of the covalent complex formed between E. coli topoisomerase I core domains and cleaved DNA (PDB 3PX7) [60]. Earlier studies implicated these enzymes in a variety of processes in both prokaryotes and eukaryotes, including DNA replication, transcription, recombination, and chromosome segregation. Rodrguez A.C., Stock D. Crystal structure of reverse gyrase: Insights into the positive supercoiling of DNA. Cuya SM, Bjornsti MA, van Waardenburg RCAM. Accessibility Studies performed over the past 3 years have provided new insight into the roles of various topoisomerases in maintaining eukaryotic chromosome structure and facilitating the decatenation of daughter chromosomes at cell division. The figure is modified from a published version for potential TOP3B action on mRNA in [29]. Human topoisomerases and their roles in genome stability and - Nature Topoisomerase 3 Is Required for Decatenation and Segregation of Human mtDNA. Domain structure of topoisomerase 11 enzymes from prokaryotes and eukaryotes A schematic representation of the domain . Recent Advances in Chemotherapeutics for Leishmaniasis: Importance of the Cellular Biochemistry of the Parasite and Its Molecular Interaction with the Host. Nikolov D.B., Burley S.K. Sequence selectivity of RNA cleavage by type IA topoisomerases have not been analyzed. Content may be subject to copyright. A water molecule has been postulated to be acting as the general base for deprotonation of the tyrosine hydroxyl nucleophile for DNA cleavage [57,62]. Characterization of molecular interactions between Escherichia coli RNA polymerase and topoisomerase I by molecular simulations. On the contrary, most eukaryotes utilize type I topoisomerases, that cut a single strand of DNA, during the movement of the replication fork.Type I Topoisomerases (creates s. MeSH Yang Y., McBride K.M., Hensley S., Lu Y., Chedin F., Bedford M.T. Transcription incurs topological constraints that result from the progression, Figure 3 |. Structural and mechanistic insight into Holliday-junction dissolution by Topoisomerase III and RMI1. Inclusion in an NLM database does not imply endorsement of, or agreement with, Studies of bacterial topoisomerases I and III at the single-molecule level. In these crystal structures, a tyrosine side chain (Tyr177 in E. coli topoisomerase I) wedges between the 4 and 5 DNA bases and creates a kink in the G-strand. Keywords: 8600 Rockville Pike According to structural homologies and reaction mechanisms, type II topoisomerases can be subdivided into two subgroups: type IIA (including gyrase and topoisomerase IV found in prokaryotes; topoisomerase II and topoisomerase II in humans) and type IIB (topoisomerase VI and VIII found in archaea and bacteria) [39,40,41]. Kirkegaard K., Wang J.C. Bacterial DNA topoisomerase I can relax positively supercoiled DNA containing a single-stranded loop. Federal government websites often end in .gov or .mil. 2017 Oct 27;429(21):3156-3167. doi: 10.1016/j.jmb.2016.08.005. Annu Rev Biochem. Moreover, the presence of divalent ions cannot compensate for the loss of this arginine residue in G-strand religation, resulting in a dominant lethal cell killing because of the accumulation of topoisomerase I-mediated DNA breaks [76]. DNA supercoiling; DNA-activated protease; DNA-protein crosslink; chromatin organization; topoisomerase poison. A thermostable enzyme with both bacterial and eucaryal features. The D111N mutation of E. coli topoisomerase I was found to be extremely lethal because the resulting deficiency in DNA religation leads to the accumulation of the covalent intermediate [59]. The .gov means its official. Post-translational regulation of Tyrosyl-DNA phosphodiesterase (TDP1 and TDP2) for the repair of the trapped topoisomerase-DNA covalent complex. The Zn(II) Binding Motifs of E. coli DNA Topoisomerase I Is Part of a High-Affinity DNA Binding Domain. Epub 2021 Sep 6. The .gov means its official. Mammalian Tyrosyl-DNA Phosphodiesterases in the Context of Mitochondrial DNA Repair. Here we review recent developments and identify further questions raised by these new findings. Type II topoisomerase - Wikipedia A conformational change in the protein then allows the intact complementary strand to be passed through the protein-linked break, followed by religation of the cleaved DNA. In addition to the relaxation of negatively supercoiled DNA and preventing hypernegative supercoiling, type IA topoisomerases can also catalyze the decatenation of replication intermediates [15,16,17,18,19] and the knotting or unknotting of single-stranded DNA circles or nicked duplex DNA [20,21]. Type IA topoisomerase introduces a transient cleavage of the single DNA strand and forms a covalent linkage to the 5-terminal phosphate of the cleaved DNA, followed by the passing of an intact strand (T-strand) of DNA through the DNA break in the cleaved G-strand and the subsequent religation of the nicked G-strand of DNA. MeSH Narula G., Annamalai T., Aedo S., Cheng B., Sorokin E., Wong A., Tse-Dinh Y.-C. Importantly, such trapped TOP1-DNA covalent complexes, formed either during ribonucleotide removal or as a consequence of drug action, activate several repair processes, including processes involving the recently described nuclear proteases SPARTAN and GCNA-1. sharing sensitive information, make sure youre on a federal In the structure of the E. coli topoisomerase I covalent complex and M. tuberculosis topoisomerase I noncovalent complex representing the pretransition state (PDB 6CQ1), this cytosine base fits into a sterically restrictive cavity formed by residues that are conserved in the topoisomerase I and reverse gyrase sequences (highlighted in blue in Figure 3D). Relaxation of hypernegatively supercoiled DNA. Overexpression of the D111N mutant topoisomerase I is toxic to the cells because of deficiency in DNA religation resulting in the accumulation of the cleavage complex on chromosomal DNA. An enzyme called helicase unwinds the DNA by breaking the hydrogen bonds between the nitrogenous base pairs. (October 2021) There are two subclasses of type II topoisomerases, type IIA and IIB. DNA topoisomerases are enzymes that catalyze changes in the torsional and flexural strain of DNA molecules. Extensive free-energy simulations identify water as the base in nucleotide addition by DNA polymerase. Federal government websites often end in .gov or .mil. A single Mg2+ is bound directly to the Glu side chain of the TOPRIM motif and through a water molecule to the first Asp of the TOPRIM DxD. DNA topoisomerase-targeting chemotherapeutics: what's new? Mutation of the TOPRIM glutamate to alanine or glutamine abolished the DNA cleavage and relaxation activity [62] but did not affect the RNA cleavage activity observed for M. smegmatis topoisomerase I, suggesting that the 2-OH of RNA could potentially participate in the proton relay for the RNA cleavage [30]. In bacteria, topoisomerase I belonging to the type IA subfamily relaxes the negative supercoiling generated by transcription behind the RNA polymerase complex to prevent the formation of DNARNA hybrids/R-loops that in turn can inhibit the transcription process [6,10,12,13,14]. sharing sensitive information, make sure youre on a federal Mondragn A., DiGate R. The structure of Escherichia coli DNA topoisomerase III. Mt: Mycobacterium tuberculosis, Sm: Streptococcus mutans, Af: Archaeoglobus fulgidus, Sc: Saccharomyces cerevisiae, Hs: Homo sapiens. 8600 Rockville Pike Bethesda, MD 20894, Web Policies Genome circularization can be mediated by RNARNA interactions and proteins binding to the 5 and 3 ends. Nat Rev Mol Cell Biol. Narula G., Tse-Dinh Y.-C. Residues of E. coli topoisomerase I conserved for interaction with a specific cytosine base to facilitate DNA cleavage. Mechanisms of DNA replication termination - Nature The number of DNA polymerases in eukaryotes is much more than prokaryotes: 14 are known, of which five are known to have major roles during replication and have been well studied. Examples of type IA topoisomerases include prokaryotic topo I and III, eukaryotic topo III and III and the archaeal enzyme reverse gyrase. PMC The guiding of the T-strand in and out of the toroid hole is also not well understood. The mechanism of the G-strand gate opening and T-strand transport by type IA topoisomerases remains to be fully elucidated. Introduction Normal cell growth requires replication of the genome and regulated transcription. This site needs JavaScript to work properly. (C) Alignment of residues strictly conserved at the active site and type IA topoisomerases including bacterial topoisomerase I, reverse gyrase (RG) and topoisomerase III from prokaryotes and eukaryotes. Epub 2016 Aug 7. Li Y, Zhao D, Zhang W, Yang M, Wu Z, Shi W, Lan S, Guo Z, Yu H, Wu D. J Transl Med. National Library of Medicine The https:// ensures that you are connecting to the The proteinprotein interactions with other cellular proteins are relevant for the physiological functions and regulation of type IA topoisomerase activities [38]. The positively charged side chain of the strictly conserved arginine residue separated by one residue from the active site tyrosine (Arg321 in E. coli topoisomerase I, Figure 3C) plays a critical role in stabilizing the negative charge on the hydroxyl nucleophile and the transition state. The PubMed wordmark and PubMed logo are registered trademarks of the U.S. Department of Health and Human Services (HHS). GCNA Interacts with Spartan and Topoisomerase II to Regulate Genome Stability. This supercoiling increases with the progression of the replication fork. Bocquet N., Bizard A.H., Abdulrahman W., Larsen N.B., Faty M., Cavadini S., Bunker R.D., Kowalczykowski S.C., Cejka P., Hickson I.D., et al. Eukaryotic DNA polymerases in DNA replication and DNA repair. Here, we demonstrate that in mitosis, spindles induce a potent positive supercoiling activity on plasmid DNA. 2023 Jun 23;14(1):3762. doi: 10.1038/s41467-023-39374-9. Crystal structure of a covalent intermediate in DNA cleavage and rejoining by Escherichia coli DNA topoisomerase I. Tan K., Zhou Q., Cheng B., Zhang Z., Joachimiak A., Tse-Dinh Y.-C. DNA topoisomerases: structure, function, and mechanism. Wu H.-Y., Shyy S., Wang J.C., Liu L.F. Chen S.H., Chan N.-L., Hsieh T.-S. New Mechanistic and Functional Insights into DNA Topoisomerases. TnpB may also be the ancestor of the eukaryotic transposon-encoded Fanzor (Fz) proteins 4,7, raising the possibility that eukaryotes are also equipped with CRISPR-Cas/OMEGA-like programmable RNA . Huang L., Wang Z., Narayanan N., Yang Y. Arginine methylation of the C-terminus RGG motif promotes TOP3B topoisomerase activity and stress granule localization. The D111N mutation in the E. coli topoisomerase I N-terminal core domain fragment enabled the isolation of the covalent complex formed with cleaved oligonucleotide for structural determination [60]. Careers. In the topoisomerase I cleavage complex (TOP1cc) (top), the 3 DNA end is covalently linked to the active site tyrosine (Y). Michael Blaber Florida State University Unwinding of the helix during DNA replication (by the action of helicase) results in supercoiling of the DNA ahead of the replication fork. Bizard A.H., Yang X., Dbat H., Fogg J.M., Zechiedrich L., Strick T.R., Garnier F., Nadal M. TopA, the Sulfolobus solfataricus topoisomerase III, is a decatenase. Bookshelf The site is secure. . Domain arrangement in E. coli topoisomerase I as seen in the crystal structure of the N-terminal core domains (PDB 1ECL) or full-length enzyme (PDB 4RUL). The amino acid sequences of the 12 eukaryotic type I topoisomerases listed in Table I were aligned using the Pile-Up function of the GCG program (Genetics Computer Group, Inc.). Topoisomerase- Definition, Types, Structure, Functions, Mechanism Certain enzymes play crucial roles in these vital cellular processes. Siaw G.E.-L., Liu I.-F., Lin P.-Y., Been M.D., Hsieh T.-S. DNA and RNA topoisomerase activities of Top3 are promoted by mediator protein Tudor domain-containing protein 3. Domain structure of topoisomerase 11 enzymes from prokaryotes and Ribonucleotides and Transcription-Associated Mutagenesis in Yeast. For example, DNA polymerase enzymes are essential for DNA replication [1] and RNA polymerases are required for transcription [2]. Flexible loops connect between these C-terminal domains and to the N-terminal core domains. doi: 10.15252/embr.202256458. Aravind L., Leipe D.D., Koonin E.V. Careers, Unable to load your collection due to an error. Distinct actions of topoisomerase poisons and inhibitors. Crystal structure of the 65-kilodalton amino-terminal fragment of DNA topoisomerase I from the gram-positive model organism Streptococcus mutans. An additional strictly conserved arginine residue in domain D4 (corresponding to Arg507 in E. coli topoisomerase I, Figure 3D) also interacts with a phosphate of the G-strand [60,62]. Positive supercoiling (Sc. The origin of replication is recognized by certain proteins that bind to this site. Here we characterize a prokaryotic counterpart to the eukaryotic topoisomerase I in the hyperthermophilic methanogen Methanopyrus kandleri. Brochu J., Breton E.V., Drolet M. Supercoiling, R-Loops, Replication and the Functions of Bacterial Type 1A Topoisomerases. Structure of a Complex between E. coli DNA Topoisomerase I and Single-Stranded DNA. Sato Y, Yoshino H, Ishikawa J, Monzen S, Yamaguchi M, Kashiwakura I. Sci Rep. 2023 Jul 4;13(1):10762. doi: 10.1038/s41598-023-37981-6. What is the difference between a prokaryotic and a eukaryotic - Quora Competing interests: Dr. Kaufmann indicates that he is the named co-inventor on a patent held by Mayo Clinic regarding the use of antibodies to TOPccs as theranostic reagents. The DNA topoisomerases are ubiquitous enzymes that fulfil vital roles in the replication, transcription and recombination of DNA by carrying out DNA-strand passage reactions. Accessibility Topoisomerase inhibitor - Wikipedia Narula G., Becker J., Cheng B., Dani N., Abrenica M.V., Tse-Dinh Y.-C. In addition, recent studies have demonstrated that the incorporation of ribonucleotides into DNA results in trapping of topoisomerase I (TOP1)-DNA covalent complexes during aborted ribonucleotide removal. Each strand in the double helix acts as a template for synthesis of a new, complementary strand. Other mutations that affect Mg2+ binding at the active site of bacterial topoisomerase I also resulted in bacterial cell death when the mutant topoisomerase I was overexpressed because of the inhibition of DNA religation and accumulation of DNA breaks [79,80,81,82]. All tangled up: How cells direct, manage and exploit topoisomerase function. Epub 2020 Mar 7. However, when Arg321 in E. coli topoisomerase I or the corresponding Arg327 in Yersinia pestis topoisomerase I was substituted with an aromatic residue, DNA cleavage could still take place but became divalent ion dependent [76]. An official website of the United States government. Such a distance was estimated by molecular dynamics simulation to be required for the passage of a double-stranded DNA through the break for a catenation/decatenation reaction involving double-stranded circular DNA with a nick or single-stranded gap [110]. Before replication can start, the DNA . In addition to acting as a ligand for Mg2+, the TOPRIM glutamate side chain has been proposed to interact directly during DNA cleavage as a general acid [57] with the G-strand 3-hydroxyl leaving group to provide a proton from a nearby positively charged histidine (His365 in E. coli topoisomerase I) side chain via proton relay through the D111 side chain [62,83]. Tiwari P.B., Chapagain P.P., Banda S., Darici Y., Uren A., Tse-Dinh Y.-C. Resistance to Antibody-Drug Conjugates Targeting HER2 in Breast Cancer: Molecular Landscape and Future Challenges. Tan K., Cao N., Cheng B., Joachimiak A., Tse-Dinh Y.-C. In the future, it will be important to understand the differential requirements for type II topoisomerases in eukaryotic and prokaryotic systems. Nat Rev Mol Cell Biol. These enzymes span all domains of life and are essential for function. No competing interests were disclosed.No competing interests were disclosed.No competing interests were disclosed. The RGG-box in topoisomerase III may contribute to both DNA and RNA binding. Accessibility 2016;8(3):221231. Many other type IA topoisomerases in all three domains of life have been found to possess RNA topoisomerase activity [28,29,30]. The deletion of the RGG-box from topoisomerase III reduced both the DNA and RNA topoisomerase activities [24]. Biochemistry, DNA Replication - StatPearls - NCBI Bookshelf ), 2Biomolecular Sciences Institute, Florida International University, Miami, FL 33199, USA, 3Biochemistry PhD Program, Florida International University, Miami, FL 33199, USA. The essential steps of replication are the same as in prokaryotes. (C) When a translating ribosome or a helicase unwinds a duplex region in a viral RNA hairpin, and if the hairpin is bound to an immobile RNP or cellular matrix, the helical torsion will need to be relaxed by TOP3B. These two enzymes are now classied in differentgroups: Type IA for the prokaryotic enzyme and type IB for the eukaryoticenzyme (see Section 5.2). It was the type I topoisomerase. Figure 4 |. Type IA topoisomerases, ubiquitous in bacteria, archaea and eukarya [41], can relax negatively supercoiled DNA but not positive supercoils because of the requirement of single-stranded DNA for binding [42]. Ceramella J, Troiano R, Iacopetta D, Mariconda A, Pellegrino M, Catalano A, Saturnino C, Aquaro S, Sinicropi MS, Longo P. Antibiotics (Basel). Fouque K.J.D., Garabedian A., Leng F., Tse-Dinh Y.-C., Fernandez-Lima F. Microheterogeneity of Topoisomerase IA/IB and Their DNA-Bound States. The G-strand of DNA fits into a binding groove in domain D4, and follows the path of one strand of a B form DNA, as observed in the subsequently obtained cocrystals of E. coli topoisomerase III (PDB 1I7D) [57] and topoisomerase I (PDB 1MW8) [58]. Cao N., Tan K., Zuo X., Annamalai T., Tse-Dinh Y.-C. Mechanistic insights from structure of Mycobacterium smegmatis topoisomerase I with ssDNA bound to both N- and C-terminal domains. HHS Vulnerability Disclosure, Help ToprimA conserved catalytic domain in type IA and II topoisomerases, DnaG-type primases, OLD family nucleases and RecR proteins. Epub 2021 Sep 6. ; graphics preparation, S.F., T.D., Y.-C.T.-D.; writingreview and editing, S.F., Y.-C.T.-D. All authors have read and agreed to the published version of the manuscript. Figure 1 |. A Mg2+ ion (in green) interacts with the scissile phosphate and TOPRIM acidic residues. Brown P.O., Cozzarelli N.R. In certain species of bacteria in the Actinobacteria phylum including members of the Mycobacterium and Streptomyces genera, the C-terminal domains are organized with repeated Topo_C_Rpt domains first predicted based on the M. tuberculosis topoisomerase I crystal structure [50] that do not have Zn(II)-binding cysteines, ending with a tail rich in positively charged lysines and arginines. Unable to load your collection due to an error, Unable to load your delegates due to an error. government site. An orthogonal single-molecule experiment reveals multiple-attempt dynamics of type IA topoisomerases. DNA gyrase, or simply gyrase, is an enzyme within the class of topoisomerase and is a subclass of Type II topoisomerases [1] that reduces topological strain in an ATP dependent manner while double-stranded DNA is being unwound by elongating RNA-polymerase [2] or by helicase in front of the progressing replication fork. Type I topoisomerases can be divided into type IA and type IB, IC subgroups [41]. Normal cell growth requires replication of the genome and regulated transcription. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (, topoisomerase, type IA, DNA supercoiling, RNA topology, decatenation. Deletion of TOP3, a component of FMRP-containing mRNPs, contributes to neurodevelopmental disorders. Stoll G., Pietilinen O.P.H., Linder B., Suvisaari J., Brosi C., Hennah W., Leppa V., Torniainen M., Ripatti S., Ala-Mello S., et al. Molecular mechanism of DNA replication (article) | Khan Academy Drolet M. Growth inhibition mediated by excess negative supercoiling: The interplay between transcription elongation, R-loop formation and DNA topology. 2001;70:369413. The ability to unknot single-stranded RNA circles was first observed for Escherichia coli topoisomerase III [22]. 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) topoisomerases. Overall, we discover fundamental differences in transcriptional control by DNA supercoiling between bacteria and yeast and show that rapid supercoiling release in eukaryotes ensures proper gene expression of neighboring genes. Stewart L., Redinbo M.R., Qiu X., Hol W.G.J., Champoux J.J. A Model for the Mechanism of Human Topoisomerase I. Baker N.M., Rajan R., Mondragn A. P30 CA013148/CA/NCI NIH HHS/United States, P50 CA136393/CA/NCI NIH HHS/United States, R01 CA190423/CA/NCI NIH HHS/United States, NCI CPTC Antibody Characterization Program, Champoux JJ: DNA topoisomerases: structure, function, and mechanism. RNA polymerase II transcription initiation: A structuralview. Role of RNA-binding proteins during the late stages of Flavivirus replication cycle. Type II DNA topoisomerase activity is required to change DNA topology. In the structure of reverse gyrases from Archaeoglobus fulgidus (PDB 1GKU) [55] and T. maritima (PDB 4DDU) [56], type IA core domains are found in the C-terminal region preceded by the helicase-like domain [55]. Type I topoisomerase - Wikipedia The Topo_C_Rpt fold has an antiparallel four-stranded -sheet flanked by a C-terminal helix on one side (Figure 4A). 2021 Sep;60:234-244. doi: 10.1016/j.mito.2021.08.017. (A) Active site of the M. tuberculosis topoisomerase I noncovalent complex (PDB 6CQ2) showing an interaction between the catalytic tyrosine and adjacent arginine with the scissile phosphate of the G-strand (in gold). Flexibility at Gly-194 Is Required for DNA Cleavage and Relaxation Activity ofEscherichia coliDNA Topoisomerase I. Cheng B., Feng J., Mulay V., Gadgil S., Tse-Dinh Y.-C. Site-directed Mutagenesis of Residues Involved in G Strand DNA Binding byEscherichia coliDNA Topoisomerase I. Bachar A., Itzhaki E., Gleizer S., Shamshoom M., Milo R., Antonovsky N. Point mutations in topoisomerase I alter the mutation spectrum in E. coli and impact the emergence of drug resistance genotypes. Perry K., Mondragn A. Biochemical Characterization of an Invariant Histidine Involved inEscherichia coliDNA Topoisomerase I Catalysis. (i) Apo enzyme; (ii) C-terminal domains (green) bind ssDNA as T-strand (red); (iii) ssDNA or G-strand (yellow) binds the N-terminal domains (blue); (iv) Active site tyrosine (red circle) becomes accessible; (v) Cleavage of the G-strand and gate opening; (vi) Passage of T-strand inside the toroid; (vii) Gate closing and trapping of T-strand; (viii) Religation of the G-strand; (ix) Gate opening and release of dsDNA. Godin K.S., Varani G. How Arginine-Rich Domains Coordinate mRNA Maturation Events. FOIA http://creativecommons.org/licenses/by/4.0/. This review on the mechanism of type IA topoisomerases will focus more on the results from recent structural, biophysical, biochemical and genetic studies that help us gain a better understanding of how type IA topoisomerases can act as magicians to manipulate the topology of both DNA and RNA, in addition to key remaining questions on the catalytic mechanism. 2011; Forterre 2012; Pommier et al. Biophys Rev. [5] This is brought about by the movement of an alpha helix that follows a strictly conserved glycine residue (Gly194 in E. coli topoisomerase I, Figure 3D). Zhu C.X., Samuel M., Pound A., Ahumada A., Tse-Dinh Y.C. Tetracysteine motifs that form the zinc ribbon fold [88] can be found in the topoisomerase I gene of a majority of bacterial species. DNA topoisomerases are ubiquitous enzymes that are essential for solving topological problems inherent to the helical structure of DNA ( Champoux 2001; Forterre and Gadelle 2009; Wang 2009; Vos et al. government site. An official website of the United States government. Type IA and type IIA topoisomerases require divalent ions for catalytic activity. Type IA topoisomerases have since evolved to catalyze the resolution of topological barriers encountered by genomes that require the passing of nucleic acid strand(s) through a break on a single DNA or RNA strand. 2016 ). Forterre P., Gadelle D. Phylogenomics of DNA topoisomerases: Their origin and putative roles in the emergence of modern organisms. It is expected that upon binding of the G-strand at the active site, the scissile phosphate will displace the water molecules seen in the crystal structure as ligands for the Mg2+. 10.1038/nrm831 2023 Feb 10;15(4):1130. doi: 10.3390/cancers15041130. Copyright: 2019 Bjornsti MA and Kaufmann SH. Active site residues assist in the nucleophilic attack of a phosphodiester bond between two nucleotides to form a covalent intermediate with a 5-phosphotyrosine linkage to the cleaved nucleic acid. Site-directed mutagenesis of E. coli topoisomerase at Gly194, Arg195 and Gln197 confirmed that mutations at these residues reduced the relaxation activity significantly [63,64]. Moreover, wild-type yeast minimizes supercoiling-mediated inhibition by maintaining sufficient levels of topoisomerases. Wang H., Di Gate R.J., Seeman N.C. An RNA topoisomerase. DNA topoisomerase V is a relative of eukaryotic topoisomerase - PubMed Direct evidence and characterization of additional transient intermediates that are part of the proposed catalytic cycle would further our understanding of the mechanism at the molecular level. Kondekar S.M., Gunjal G.V., Radicella J.P., Rao D.N. The key difference between prokaryotic and eukaryotic cells is that eukaryotic cells have a membrane-bound nucleus (and membrane-bound organelles), whereas prokaryotic cells lack a nucleus. Ma J., Wang M.D. Okazaki fragments are short sequences of DNA nucleotides (approximately 150 to 200 base pairs long in eukaryotes) which are synthesized discontinuously and later linked together by the enzyme DNA ligase to create the lagging strand during DNA replication. Pharmaceuticals (Basel).

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