Topoisomerase II Research Articles

Structural and functional insights into the T-even type bacteriophage topoisomerase II.

T-even type bacteriophages are virulent phages commonly used as model organisms, playing a crucial role in understanding various biological processes. One such process involves the regulation of DNA topology during phage replication upon host infection, governed by type IIA DNA topoisomerases. In spite of various studies on prokaryotic and eukaryotic counterparts, viral topoisomerase II remains insufficiently understood, especially the unique domain composition of T4 phage. In this study, we determine the cryo-EM structures of topoisomerase II from T4 and T6 phages, including full-length structures of both apo and DNA-binding states which have never been determined before. Together with other conformational states, these structures provide an explicit blueprint of mechanisms of phage topoisomerase II. Particularly, the asymmetric dimeric interactions observed in cryo-EM structures of T6 phage topoisomerase II ATPase domain and central domain bound with DNA shed light on the asynchronous ATP usage and asynchronous cleavage of the G-segment DNA, respectively. The elucidation of phage topoisomerase II's structures and functions not only enhances our understanding of mechanisms and evolutionary parallels with prokaryotic and eukaryotic homologs but also highlights its potential as a model for developing type IIA topoisomerase inhibitors.

Structural insights into the assembly of type IIA topoisomerase DNA cleavage-religation center.

The ability to catalyze reversible DNA cleavage and religation is central to topoisomerases' role in regulating DNA topology. In type IIA topoisomerases (Top2), the formation of its DNA cleavage-religation center is driven by DNA-binding-induced structural rearrangements. These changes optimally position key catalytic modules, such as the active site tyrosine of the WHD domain and metal ion(s) chelated by the TOPRIM domain, around the scissile phosphodiester bond to perform reversible transesterification. To understand this assembly process in detail, we report the catalytic core structures of human Top2α and Top2β in an on-pathway conformational state. This state features an in trans formation of an interface between the Tower and opposing TOPRIM domain, revealing a groove for accommodating incoming G-segment DNA. Structural superimposition further unveils how subsequent DNA-binding-induced disengagement of the TOPRIM and Tower domains allows a firm grasp of the bound DNA for cleavage/religation. Notably, we identified a previously undocumented protein-DNA interaction, formed between an arginine-capped C-terminus of an α-helix in the TOPRIM domain and the DNA backbone, significantly contributing to Top2 function. This work uncovers a previously unrecognized role of the Tower domain, highlighting its involvement in anchoring and releasing the TOPRIM domain, thus priming Top2 for DNA binding and cleavage.

Rapid, DNA-induced interface swapping by DNA gyrase

DNA gyrase, a ubiquitous bacterial enzyme, is a type IIA topoisomerase formed by heterotetramerisation of 2 GyrA subunits and 2 GyrB subunits, to form the active complex. DNA gyrase can loop DNA around the C-terminal domains (CTDs) of GyrA and pass one DNA duplex through a transient double-strand break (DSB) established in another duplex. This results in the conversion from a positive (+1) to a negative (–1) supercoil, thereby introducing negative supercoiling into the bacterial genome by steps of 2, an activity essential for DNA replication and transcription. The strong protein interface in the GyrA dimer must be broken to allow passage of the transported DNA segment and it is generally assumed that the interface is usually stable and only opens when DNA is transported, to prevent the introduction of deleterious DSBs in the genome. In this paper, we show that DNA gyrase can exchange its DNA-cleaving interfaces between two active heterotetramers. This so-called interface ‘swapping’ (IS) can occur within a few minutes in solution. We also show that bending of DNA by gyrase is essential for cleavage but not for DNA binding per se and favors IS. Interface swapping is also favored by DNA wrapping and an excess of GyrB. We suggest that proximity, promoted by GyrB oligomerization and binding and wrapping along a length of DNA, between two heterotetramers favors rapid interface swapping. This swapping does not require ATP, occurs in the presence of fluoroquinolones, and raises the possibility of non-homologous recombination solely through gyrase activity. The ability of gyrase to undergo interface swapping explains how gyrase heterodimers, containing a single active-site tyrosine, can carry out double-strand passage reactions and therefore suggests an alternative explanation to the recently proposed ‘swivelling’ mechanism for DNA gyrase (Gubaev et al., 2016).

Rapid, DNA-induced interface swapping by DNA gyrase.

DNA gyrase, a ubiquitous bacterial enzyme, is a type IIA topoisomerase formed by heterotetramerisation of 2 GyrA subunits and 2 GyrB subunits, to form the active complex. DNA gyrase can loop DNA around the C-terminal domains (CTDs) of GyrA and pass one DNA duplex through a transient double-strand break (DSB) established in another duplex. This results in the conversion from a positive (+1) to a negative (-1) supercoil, thereby introducing negative supercoiling into the bacterial genome by steps of 2, an activity essential for DNA replication and transcription. The strong protein interface in the GyrA dimer must be broken to allow passage of the transported DNA segment and it is generally assumed that the interface is usually stable and only opens when DNA is transported, to prevent the introduction of deleterious DSBs in the genome. In this paper, we show that DNA gyrase can exchange its DNA-cleaving interfaces between two active heterotetramers. This so-called interface 'swapping' (IS) can occur within a few minutes in solution. We also show that bending of DNA by gyrase is essential for cleavage but not for DNA binding per se and favors IS. Interface swapping is also favored by DNA wrapping and an excess of GyrB. We suggest that proximity, promoted by GyrB oligomerization and binding and wrapping along a length of DNA, between two heterotetramers favors rapid interface swapping. This swapping does not require ATP, occurs in the presence of fluoroquinolones, and raises the possibility of non-homologous recombination solely through gyrase activity. The ability of gyrase to undergo interface swapping explains how gyrase heterodimers, containing a single active-site tyrosine, can carry out double-strand passage reactions and therefore suggests an alternative explanation to the recently proposed 'swivelling' mechanism for DNA gyrase (Gubaev et al., 2016).

Molecular choreography: Unveiling the dynamic landscape of type IIA DNA topoisomerases before T-segment passage through all-atom simulations

Type IIA DNA topoisomerases are molecular nanomachines responsible for controlling topological states of DNA molecules. Here, we explore the dynamic landscape of yeast topoisomerase IIA during key stages of its catalytic cycle, focusing in particular on the events preceding the passage of the T-segment. To this end, we generated six configurations of fully catalytic yeast topo IIA, strategically inserted a T-segment into the N-gate in relevant configurations, and performed all-atom simulations.The essential motion of topo IIA protein dimer was characterized by rotational gyrating-like movement together with sliding motion within the DNA-gate. Both appear to be inherent properties of the enzyme and an inbuilt feature that allows passage of the T-segment through the cleaved G-segment. Coupled dynamics of the N-gate and DNA-gate residues may be particularly important for controlled and smooth passage of the T-segment and consequently the prevention of DNA double-strand breaks. QTK loop residue Lys367, which interacts with ATP and ADP molecules, is involved in regulating the size and stability of the N-gate. The unveiled features of the simulated configurations provide insights into the catalytic cycle of type IIA topoisomerases and elucidate the molecular choreography governing their ability to modulate the topological states of DNA topology.

Synthesis, XRD, spectroscopic (UV–Vis, IR, EPR) and biological evaluations of cobalt(II)-ciprofloxacin complex as antimicrobial agent: In silico molecular docking and ADME

Cobalt(II)-Ciprofloxacin complex ([Co(Cip)2(H2O)2].2(H2PO4).8(H2O); Cip=Ciprofloxacin) containing phosphoric acid was synthesized and its structure was characterized by numerous analytical techniques such as FT-IR (Fourier Transform Infrared), UV–Vis (Ultraviolet-Visible), EPR (Electron Paramagnetic Resonance), TGA (Thermogravimetric Analysis), elemental analysis and SCXRD (single crystal X-ray diffraction) for structural elucidation. According to XRD data, the environment of Cobalt has octahedral geometry with ciprofloxacin ligand bonded as bidentate (keto and carboxyl group) and aqua ligand. It was revealed that the metal complex left metal oxide and other residues as the final product in the multi-step decomposition TGA curve in the range of 20–1000 °C. The contribution of intermolecular interactions that lead to molecular packing was analyzed using Hirshfeld surface analysis. The synthesized complex was tested for antimicrobial activity against Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Candida albicans. We conducted a molecular docking study using AUTODOCK 4.2 to investigate the binding energy and interaction modes of a highly potent microbial complex with three different enzymes: S. aureus DNA gyrase (PDB ID: 2XCT), GyrA (PDB ID: 3LPX), and mycobacterium tuberculosis gyrase type IIA topoisomerase (PDB ID: 3UC1).

BLM and BRCA1-BARD1 coordinate complementary mechanisms of joint DNA molecule resolution

The Bloom syndrome helicase BLM interacts with topoisomerase IIIα (TOP3A), RMI1, and RMI2 to form the BTR complex, which dissolves double Holliday junctions and DNA replication intermediates to promote sister chromatid disjunction before cell division. In its absence, structure-specific nucleases like the SMX complex (comprising SLX1-SLX4, MUS81-EME1, and XPF-ERCC1) can cleave joint DNA molecules instead, but cells deficient in both BTR and SMX are not viable. Here, we identify a negative genetic interaction between BLM loss and deficiency in the BRCA1-BARD1 tumor suppressor complex. We show that this is due to a previously overlooked role for BARD1 in recruiting SLX4 to resolve DNA intermediates left unprocessed by BLM in the preceding interphase. Consequently, cells with defective BLM and BRCA1-BARD1 accumulate catastrophic levels of chromosome breakage and micronucleation, leading to cell death. Thus, we reveal mechanistic insights into SLX4 recruitment to DNA lesions, with potential clinical implications for treating BRCA1-deficient tumors.

In Silico Activity Prediction and Docking Studies of the Binding Mechanisms of Levofloxacin Structure Derivatives to Active Receptor Sites of Bacterial Type IIA Topoisomerases

The need for new antimicrobial agents (AntAg) is driven by the persistent antibiotic resistance in microorganisms, as well as the increasing frequency of pandemics. Due to the deficiency of AntAg, research aimed at developing speedy approaches to find new drug candidates is relevant. This study aims to conduct an in silico study of the biological activity spectrum as well as the molecular binding mechanisms of four structurally different forms of levofloxacin (Lvf) with bacterial topoisomerases targets of type IIA (DNA gyrase and topoisomerase IV) to enable the development of drugs with an improved characterization of the safety profile. To achieve this goal, a number of software products were used, such as ChemicPen v. 2.6, PyMol 2.5, Avogadro 1.2.0, PASS, AutoDockTools 1.5.7 with the new generation software Autodock Vina. These software products are the first to be made available for visualization of clusters with determination of ligand-receptor pair binding affinity, as well as clustering coordinates and proposed mechanisms of action. One of the real structures of Lvf, a decarboxylated derivative, was obtained with tribochemical (TrbCh) exposure. The action spectrum of molecular ligands is described based on a Bayesian probability activity prediction model (PASS software Version 2.0). Predicted and real (PMS and RMS) molecular structures of Lvf, with decreasing levels of structural complexity, were translated into descriptors via Wiener (W), Balaban (Vs), Detour (Ip), and Electropy € indices. The 2D «structure-activity» diagrams were used to differentiate closely related structures of levofloxacin. PMS and RMS were visualized as 3D models of the ligand-receptor complexes. The contact regions of RMS and PMS with key amino acid residues—SER-79, DT-15, DG-1, DA-1—were demonstrated. The intra- and inter-molecular binding sites, data on free energy (affinity values, kcal/mol), the binding constant Kb (M−1), and the number of clusters are presented. The research results obtained from the presented in silico approach to explore the spectrum of action find quantitative “structure-activity” correlations, and predict molecular mechanisms may be of applied interest for directed drug discovery.

4-aminoantipyrine linked bis-1,2,3-triazole based probes for Cu(II) sensing

Herein, two molecular probes of 4-aminoantipyrine-linked bis-1,2,3-triazoles have been studied for metal ion sensing applications. The crystal structure of one of the probes named as 4,4′-((4,4′-(((1,5-dimethyl- 3- oxo- 2- phenyl- 2,3- dihydro- 1H- pyrazol- 4-yl) azanediyl) bis(methylene))bis(1H-1,2,3-triazole-4,1-diyl))bis(methylene))dibenzonitrile (3b) crystalizes in Monoclinic crystal system in P 1 21/n 1 space group. Both probes responded more strongly and selectively to Cu(II) than other tested metal ions (K+, Na+, Mg2+, Ba2+, Ca2+, Cr3+, Mn2+, Co2+, Zn2+, Cd2+, Ni2+, Hg2+, and Pb2+). Job's plot suggested a 1:1 stoichiometric ratio of ligand and metal ion. Both probes showed an association constant of 2.48 × 103 M–1 and 3.73 × 103 M–1 through Benesi-Hildebrand (B-H) plot. The geometries of both probes and their complexes were optimized by DFT using B3LYP/6–311 G(d,p) and B3LYP/LanL2DZ basis sets, and properties were supported by Mulliken atomic charge and Molecular electrostatic potential. Molecular docking on both probes and their complexes with Type-II topoisomerases(PDB ID: 4G0V) protein was conducted to foretastes anticancer activity.

Chromatinization modulates topoisomerase II processivity

Type IIA topoisomerases are essential DNA processing enzymes that must robustly and reliably relax DNA torsional stress. While cellular processes constantly create varying torsional stress, how this variation impacts type IIA topoisomerase function remains obscure. Using multiple single-molecule approaches, we examined the torsional dependence of eukaryotic topoisomerase II (topo II) activity on naked DNA and chromatin. We observed that topo II is ~50-fold more processive on buckled DNA than previously estimated. We further discovered that topo II relaxes supercoiled DNA prior to plectoneme formation, but with processivity reduced by ~100-fold. This relaxation decreases with diminishing torsion, consistent with topo II capturing transient DNA loops. Topo II retains high processivity on buckled chromatin (~10,000 turns) and becomes highly processive even on chromatin under low torsional stress (~1000 turns), consistent with chromatin’s predisposition to readily form DNA crossings. This work establishes that chromatin is a major stimulant of topo II function.

New developments in the fight against bacterial infections : Update on antiobiotic research, development and treatment

Infections caused by pathogens with antimicrobial resistance (AMR) pose athreat to modern healthcare and have triggered the development of comprehensive national and global action plans against the spread of AMR. These include an increasing global network with the focus on rational antibiotic use, innovative strategies on antibiotic research and development, and new therapeutic approaches in antibacterial drug research. In Europe 671,689 infections associated with AMR pathogens and 33,110 deaths directly related to AMR were counted in just 1year. Globally, resistant Staphylococcus aureus, Escherichia coli, pneumococci, Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa are the most common pathogens in the context of these deaths. Resistance to antibiotics in major drug classes such as beta-lactams and fluoroquinolones is particularly common. Strategies for overcoming the global AMR crisis address research on AMR emergence and spread, promoting campaigns for responsible antibiotic use, and improving infection prevention. The identification of new antibiotics and treatment approaches and the development of new strategies to contain the spread of AMR are essential. Newly approved substances include delafloxacin, lefamulin, and meropenem-vaborbactam. New antibiotics that are well advanced in clinical trials are aztreonam-avibactam, sulbactam-durlobactam, omadacycline, and typeII topoisomerase inhibitors. Much interest is also being shown in the development of new therapeutic approaches such as bacteriophage treatment.

Cryo-EM structures of African swine fever virus topoisomerase.

African swine fever virus (ASFV) is a highly contagious virus that causes lethal hemorrhagic diseases known as African swine fever (ASF) with a case fatality rate of 100%. There is an urgent need to develop anti-ASFV drugs. We determine the first high-resolution structures of viral topoisomerase ASFV P1192R in both the closed and open C-gate forms. P1192R shows a similar overall architecture with eukaryotic and prokaryotic type II topoisomerases, which have been successful targets of many antimicrobials and anticancer drugs, with the most similarity to yeast topo II. P1192R also exhibits differences in the details of active site configuration, which are important to enzyme activity. These two structures offer useful structural information for antiviral drug design and provide structural evidence to support that eukaryotic type IIA topoisomerase likely originated from horizontal gene transfer from the virus.

Structural basis for the assembly of the DNA cleavage-religation core of type IIA topoisomerase

Structural basis for the assembly of the DNA cleavage-religation core of type IIA topoisomerase

Computational insight of antioxidant and doxorubicin combination for effective cancer therapy

Doxorubicin (DOX) is the most effective antineoplastic agent, destroys cancer cells by interrupting cellular function. However, the serious side effects on the heart limits its utility. To curb these unwanted side effects, nutritionist recommend antioxidants use along with DOX while chemotherapy. But it was not supported by various oncologists as it can alter the toxicity of DOX towards cancer cells. Therefore, here we explored the in silico pharmacokinetics and combination effect of DOX and antioxidants on topoisomerases-II (Top-II) and cyclophilin D (Cyp-D) therapeutic targets involved in cancer proliferation and post-myocardial infarction, respectively. The molecular docking study was conducted on target proteins and DOX including most prescribed antioxidants (melatonin, N-acetylcysteine (NAC), glutathione (GSH), β-carotene and vitamin C). GSH showed effective binding potential for Top-II and Cyp-D active sites, but other considered antioxidants possess low binding affinity. The highest docked conformations were subjected to molecular dynamics (MD) simulations to understand conformer stability of DOX and GSH with Cyp-D and Top-II for 100 ns. The results revealed that ligands pose at Top-II active sites where DOX showed strong binding affinity to DNA binding pocket and GSH to a buried site. The computational data summarised and proposed the GSH and DOX combination as antagonist effects on Top-II. Conversely, the binding compactness of GSH improved due to surface fit at the active pocket of Cyp-D and completely blocking DOX binding affinity, suppress adverse reactions of post-myocardial infarction. Communicated by Ramaswamy H. Sarma

Updated Review on Clinically-Relevant Properties of Delafloxacin.

The extensive use of fluoroquinolones has been consequently accompanied by the emergence of bacterial resistance, which triggers the necessity to discover new compounds. Delafloxacin is a brand-new anionic non-zwitterionic fluoroquinolone with some structural particularities that give it attractive proprieties: high activity under acidic conditions, greater in vitro activity against Gram-positive bacteria-even those showing resistance to currently-used fluoroquinolones-and nearly equivalent affinity for both type-II topoisomerases (i.e., DNA gyrase and topoisomerase IV). During phases II and III clinical trials, delafloxacin showed non-inferiority compared to standard-of-care therapy in the treatment of acute bacterial skin and skin structure infections and community-acquired bacterial pneumonia, which resulted in its approval in 2017 by the Food and Drug Administration for indications. Thanks to its overall good tolerance, its broad-spectrum in vitro activity, and its ease of use, it could represent a promising molecule for the treatment of bacterial infections.

The role of TOP2A in immunotherapy and vasculogenic mimicry in non-small cell lung cancer and its potential mechanism

Type IIA topoisomerase (TOP2A) is significantly associated with malignant tumor development, invasion, treatment and its prognosis, and has been shown to be a therapeutic target against cancer. In contrast, the role of TOP2A in the immunotherapy of non-small cell lung cancer as well as in Vasculogenic mimicry (VM) formation and its potential mechanisms are unclear. The aim of this study was to investigate the role of TOP2A in proliferation, skeleton regulation, motility and VM production in non-small cell lung cancer and its mechanisms by using bioinformatics tools and molecular biology experiments. Subgroup analysis showed that the low-risk group had a better prognosis, while the high-risk group was positively correlated with high tumor mutational load, M1-type macrophage infiltration, immune checkpoint molecule expression, and immunotherapy efficacy. As confirmed by further clinical specimens, the presence of TOP2A and VM was significantly and positively correlated with poor prognosis. Our study established a model based on significant co-expression of TOP2A genes, which significantly correlated with mutational load and immunotherapy outcomes in patients with non-small cell lung cancer. Further mechanistic exploration suggests that TOP2A plays an important role in immunotherapy and VM formation in NSCLC through upregulation of Wnt3a and PD-L1 expression.

Replication-associated formation and repair of human topoisomerase IIIα cleavage complexes

Topoisomerase IIIα (TOP3A) belongs to the conserved Type IA family of DNA topoisomerases. Here we report that human TOP3A is associated with DNA replication forks and that a “self-trapping” TOP3A mutant (TOP3A-R364W) generates cellular TOP3A DNA cleavage complexes (TOP3Accs). We show that trapped TOP3Accs that interfere with replication, induce DNA damage and genome instability. To elucidate how TOP3Accs are repaired, we explored the role of Spartan (SPRTN), the metalloprotease associated with DNA replication, which digests proteins forming DNA-protein crosslinks (DPCs). We find that SPRTN-deficient cells show elevated TOP3Accs, whereas overexpression of SPRTN lowers cellular TOP3Accs. SPRTN is deubiquitinated and epistatic with TDP2 in response to TOP3Accs. In addition, we found that MRE11 can excise TOP3Accs, and that cell cycle determines the preference for the SPRTN-TDP2 vs. the ATM-MRE11 pathways, in S vs. G2, respectively. Our study highlights the prevalence of TOP3Accs repair mechanisms to ensure normal DNA replication.

Synthesis, crystal structure, DFT/HF, Hirshfeld surface, and molecular docking analysis of 4-(tert-butyl)-4-nitro-1,1-biphenyl

4-(tert-Butyl)-4-nitro-1,1-biphenyl has been synthesized, and its structure has been characterized by using some spectroscopic and single-crystal X-ray diffraction techniques. It crystallizes in a monoclinic crystal system with space group P21/n and unit cell parameters: a = 6.4478(3) Å, b = 9.2477(4) Å, c = 23.4572(9) Å, β = 95.114(4)°, V = 1393.11(10) Å3, Z = 4. The molecular structure has been solved by using the intrinsic phasing method. The crystal structure is stabilized by C-H···O interactions. Computational studies were performed using density functional theory (DFT) and Hartree-Fock (HF) methods. The optimized geometry obtained from DFT and HF in the gas phase was compared with solid-phase experimental data retrieved from single-crystal X-ray diffraction results. Frontier molecular orbitals, such as the HOMO/LUMO energy gap, the molecular electrostatic potential, and Mulliken atomic charges, have been investigated. The HOMO LUMO energy gap of 3.97 eV indicates that the molecule is soft and highly reactive. The Hirshfeld surface analysis and their associated fingerprint plots have been used to quantitatively validate the interactions. Further insilico molecular docking studies have been performed with the molecular target Type-II topoisomerase (PDB ID: 1JIJ) and their results suggest that 4-(tert-butyl)-4-nitro-1,1-biphenyl could be considered an anticancer drug.

The expression levels of NF-κB and IKKβ in epithelial ovarian cancer and their correlation with drug resistance-related genes MDR1, TOPOII, and ERCC1.

To explore the expression levels of nuclear factor kappa B (NF-κB) and inhibitor of nuclear factor kappa B kinase (IKKβ) in epithelial ovarian cancer and the correlation analysis with multi-drug resistance-related genes 1 (MDR1), topoisomerase II (TOPOII) and nucleotide excision repair cross complementary group 1 (ERCC1). Immunohistochemical methods were used to detect the expression levels of NF-κB and IKKβ in epithelial ovarian cancer group (50 cases), ovarian benign tumor group (30 cases), and normal ovary group (10 cases). The expression levels of NF-κB, IKKβ, MDR1, TOPOII and ERCC1 messenger ribonucleic acid (mRNA) and protein were analyzed using real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot. Student's t-test and one-way ANOVA were used for comparison of numerical data. Pearson's chi-squared and Fisher's exact tests were carried out for analysis of non-numerical data. The levels of NF-κB, IKKβ, MDR1 and ERCC1 mRNA and protein were increased (P<0.05), and the expression levels of TOPOII were decreased (P<0.05) in the epithelial ovarian cancer group compared to the normal ovary and benign ovarian tumor groups. The expression of NF-κB and IKKβ in epithelial ovarian cancer was significantly increased in patients with higher tumor stage, lower differentiation and presence of lymph node metastasis and positively correlated with MDR1 expression. NF-κB and IKKβ were negatively correlated with the expression of TOPOII and antagonized each other with TOPOII. The expression of NF-κB and IKKβ was positively correlated with the expression of MDR1, and negatively correlated with the expression of TOPOII. The correlation of NF-κB, IKKβ and resistance related genes, including MDR1, TOPOII, ERCC1, can predict the resistance of chemotherapy individuals to chemotherapy.

What's on the Other Side of the Gate: A Structural Perspective on DNA Gate Opening of Type IA and IIA DNA Topoisomerases.

DNA topoisomerases have an essential role in resolving topological problems that arise due to the double-helical structure of DNA. They can recognise DNA topology and catalyse diverse topological reactions by cutting and re-joining DNA ends. Type IA and IIA topoisomerases, which work by strand passage mechanisms, share catalytic domains for DNA binding and cleavage. Structural information has accumulated over the past decades, shedding light on the mechanisms of DNA cleavage and re-ligation. However, the structural rearrangements required for DNA-gate opening and strand transfer remain elusive, in particular for the type IA topoisomerases. In this review, we compare the structural similarities between the type IIA and type IA topoisomerases. The conformational changes that lead to the opening of the DNA-gate and strand passage, as well as allosteric regulation, are discussed, with a focus on the remaining questions about the mechanism of type IA topoisomerases.