Browse
Submit Data
Databases
API
Help

Dataset Information

24 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Guiding strand passage: DNA-induced movement of the gyrase C-terminal domains defines an early step in the supercoiling cycle.

ABSTRACT: DNA gyrase catalyzes ATP-dependent negative supercoiling of DNA in a strand passage mechanism. A double-stranded segment of DNA, the T-segment, is passed through the gap in a transiently cleaved G-segment by coordinated closing and opening of three protein interfaces in gyrase. T-segment capture is thought to be guided by the C-terminal domains of the GyrA subunit of gyrase that wrap DNA around their perimeter and cause a DNA-crossing with a positive handedness. We show here that the C-terminal domains are in a downward-facing orientation in the absence of DNA, but swing up and rotate away from the gyrase body when DNA binds. The upward movement of the C-terminal domains is an early event in the catalytic cycle of gyrase that is triggered by binding of a G-segment, and first contacts of the DNA with the C-terminal domains, and contributes to T-segment capture and subsequent strand passage.

SUBMITTER: Lanz MA

PROVIDER: S-EPMC3239214 | biostudies-literature | 2011 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Guiding strand passage: DNA-induced movement of the gyrase C-terminal domains defines an early step in the supercoiling cycle.

Lanz Martin A MA Klostermeier Dagmar D

Nucleic acids research 20110831 22

DNA gyrase catalyzes ATP-dependent negative supercoiling of DNA in a strand passage mechanism. A double-stranded segment of DNA, the T-segment, is passed through the gap in a transiently cleaved G-segment by coordinated closing and opening of three protein interfaces in gyrase. T-segment capture is thought to be guided by the C-terminal domains of the GyrA subunit of gyrase that wrap DNA around their perimeter and cause a DNA-crossing with a positive handedness. We show here that the C-terminal ...[more]

PMID: 21880594

Similar Datasets

DNA-induced narrowing of the gyrase N-gate coordinates T-segment capture and strand passage.

Project description:DNA gyrase introduces negative supercoils into DNA in an ATP-dependent reaction. DNA supercoiling is catalyzed by a strand-passage mechanism, in which a T-segment of DNA is passed through the gap in a transiently cleaved G-segment. Strand passage requires the coordinated closing and opening of three protein interfaces in gyrase, the N-gate, DNA-gate, and C-gate. We show here that DNA binding to the DNA-gate of gyrase and wrapping of DNA around the C-terminal domains of GyrA induces a narrowing of the N-gate. This half-closed state prepares capture of a T-segment in the upper cavity of gyrase. Subsequent N-gate closure upon binding of ATP then poises the reaction toward strand passage. The N-gate reopens after ATP hydrolysis, allowing for further catalytic cycles. DNA binding, cleavage, and wrapping and N-gate narrowing are intimately linked events that coordinate conformational changes at the DNA and the N-gate.

| S-EPMC3161603 | biostudies-literature

Mechanisms for defining supercoiling set point of DNA gyrase orthologs: I. A nonconserved acidic C-terminal tail modulates Escherichia coli gyrase activity.

Project description:DNA topoisomerases manage chromosome supercoiling and organization in all cells. Gyrase, a prokaryotic type IIA topoisomerase, consumes ATP to introduce negative supercoils through a strand passage mechanism. All type IIA topoisomerases employ a similar set of catalytic domains for function; however, the activity and specificity of gyrase are augmented by a specialized DNA binding and wrapping element, termed the C-terminal domain (CTD), which is appended to its GyrA subunit. We have discovered that a nonconserved, acidic tail at the extreme C terminus of the Escherichia coli GyrA CTD has a dramatic and unexpected impact on gyrase function. Removal of the CTD tail enables GyrA to introduce writhe into DNA in the absence of GyrB, an activity exhibited by other GyrA orthologs, but not by wild-type E. coli GyrA. Strikingly, a "tail-less" gyrase holoenzyme is markedly impaired for DNA supercoiling capacity, but displays normal ATPase function. Our findings reveal that the E. coli GyrA tail regulates DNA wrapping by the CTD to increase the coupling efficiency between ATP turnover and supercoiling, demonstrating that CTD functions can be fine-tuned to control gyrase activity in a highly sophisticated manner.

| S-EPMC3365713 | biostudies-literature

CryoEM structures of open dimers of gyrase A in complex with DNA illuminate mechanism of strand passage.

Project description:Gyrase is a unique type IIA topoisomerase that uses ATP hydrolysis to maintain the negatively supercoiled state of bacterial DNA. In order to perform its function, gyrase undergoes a sequence of conformational changes that consist of concerted gate openings, DNA cleavage, and DNA strand passage events. Structures where the transported DNA molecule (T-segment) is trapped by the A subunit have not been observed. Here we present the cryoEM structures of two oligomeric complexes of open gyrase A dimers and DNA. The protein subunits in these complexes were solved to 4 Å and 5.2 Å resolution. One of the complexes traps a linear DNA molecule, a putative T-segment, which interacts with the open gyrase A dimers in two states, representing steps either prior to or after passage through the DNA-gate. The structures locate the T-segment in important intermediate conformations of the catalytic cycle and provide insights into gyrase-DNA interactions and mechanism.

| S-EPMC6286129 | biostudies-literature

The acidic C-terminal tail of the GyrA subunit moderates the DNA supercoiling activity of Bacillus subtilis gyrase.

Project description:Gyrase is a type II DNA topoisomerase that introduces negative supercoils into DNA in an ATP-dependent reaction. It consists of a topoisomerase core, formed by the N-terminal domains of the two GyrA subunits and by the two GyrB subunits, that catalyzes double-stranded DNA cleavage and passage of a second double-stranded DNA through the gap in the first. The C-terminal domains (CTDs) of the GyrA subunits form a ?-pinwheel and bind DNA around their positively charged perimeter. As a result, DNA is bound as a positive supercoil that is converted into a negative supercoil by strand passage. The CTDs contain a conserved 7-amino acid motif that connects blades 1 and 6 of the ?-pinwheel and is a hallmark feature of gyrases. Deletion of this so-called GyrA-box abrogates DNA bending by the CTDs and DNA-induced narrowing of the N-gate, affects T-segment presentation, reduces the coupling of DNA binding to ATP hydrolysis, and leads to supercoiling deficiency. Recently, a severe loss of supercoiling activity of Escherichia coli gyrase upon deletion of the non-conserved acidic C-terminal tail (C-tail) of the CTDs has been reported. We show here that, in contrast to E. coli gyrase, the C-tail is a very moderate negative regulator of Bacillus subtilis gyrase activity. The C-tail reduces the degree of DNA bending by the CTDs but has no effect on DNA-induced conformational changes of gyrase that precede strand passage and reduces DNA-stimulated ATPase and DNA supercoiling activities only 2-fold. Our results are in agreement with species-specific, differential regulatory effects of the C-tail in gyrases from different organisms.

| S-EPMC4007426 | biostudies-literature

Mechanisms for defining supercoiling set point of DNA gyrase orthologs: II. The shape of the GyrA subunit C-terminal domain (CTD) is not a sole determinant for controlling supercoiling efficiency.

Project description:DNA topoisomerases are essential enzymes that can overwind, underwind, and disentangle double-helical DNA segments to maintain the topological state of chromosomes. Nearly all bacteria utilize a unique type II topoisomerase, gyrase, which actively adds negative supercoils to chromosomes using an ATP-dependent DNA strand passage mechanism; however, the specific activities of these enzymes can vary markedly from species to species. Escherichia coli gyrase is known to favor supercoiling over decatenation (Zechiedrich, E. L., Khodursky, A. B., and Cozzarelli, N. R. (1997) Genes Dev. 11, 2580-2592), whereas the opposite has been reported for Mycobacterium tuberculosis gyrase (Aubry, A., Fisher, L. M., Jarlier, V., and Cambau, E. (2006) Biochem. Biophys. Res. Commun. 348, 158-165). Here, we set out to understand the molecular basis for these differences using structural and biochemical approaches. Contrary to expectations based on phylogenetic inferences, we find that the dedicated DNA wrapping domains (the C-terminal domains) of both gyrases are highly similar, both architecturally and in their ability to introduce writhe into DNA. However, the M. tuberculosis enzyme lacks a C-terminal control element recently uncovered in E. coli gyrase (see accompanying article (Tretter, E. M., and Berger, J. M. (2012) J. Biol. Chem. 287, 18636-18644)) and turns over ATP at a much slower rate. Together, these findings demonstrate that C-terminal domain shape is not the sole regulatory determinant of gyrase activity and instead indicate that an inability to tightly couple DNA wrapping to ATP turnover is why M. tuberculosis gyrase cannot supercoil DNA to the same extent as its γ-proteobacterial counterpart. Our observations demonstrate that gyrase has been modified in multiple ways throughout evolution to fine-tune its specific catalytic properties.

| S-EPMC3365746 | biostudies-literature

Reverse gyrase has heat-protective DNA chaperone activity independent of supercoiling.

Project description:Hyperthermophilic organisms must protect their constituent macromolecules from heat-induced degradation. A general mechanism for thermoprotection of DNA in active cells is unknown. We show that reverse gyrase, the only protein that is both specific and common to all hyperthermophiles, reduces the rate of double-stranded DNA breakage approximately 8-fold at 90 degrees C. This activity does not require ATP hydrolysis and is independent of the positive supercoiling activity of the enzyme. Reverse gyrase has a minor nonspecific effect on the rate of depurination, and a major specific effect on the rate of double-strand breakage. Using electron microscopy, we show that reverse gyrase recognizes nicked DNA and recruits a protein coat to the site of damage through cooperative binding. Analogously to molecular chaperones that assist unfolded proteins, we found that reverse gyrase prevents inappropriate aggregation of denatured DNA regions and promotes correct annealing. We propose a model for a targeted protection mechanism in vivo in which reverse gyrase detects damaged DNA and acts as a molecular splint to prevent DNA breakage in the vicinity of the lesion, thus maintaining damaged DNA in a conformation that is amenable to repair.

| S-EPMC484171 | biostudies-literature

Profiling DNA supercoiling domains in vivo.

Project description:Transitions in DNA structure have the capacity to regulate genes, but have been poorly characterised in eukaryotes due to a lack of appropriate techniques. One important example is DNA supercoiling, which can directly regulate transcription initiation, elongation and coordinated expression of neighbouring genes. DNA supercoiling is the over- or under-winding of the DNA double helix, which occurs as a consequence of polymerase activity and is modulated by topoisomerase activity [5]. To map the distribution of DNA supercoiling in nuclei, we developed biotinylated 4,5,8-trimethylpsoralen (bTMP) pull-down to preferentially enrich for under-wound DNA. Here we describe in detail the experimental design, quality controls and analyses associated with the study by Naughton et al. [13] that characterised for the first time the large-scale distribution of DNA supercoiling in human cells (GEO: GSE43488 and GSE43450GSE43488GSE43450).

| S-EPMC4536041 | biostudies-literature

Carboxyl terminal domain basic amino acids of mycobacterial topoisomerase I bind DNA to promote strand passage.

Project description:Bacterial DNA topoisomerase I (topoI) carries out relaxation of negatively supercoiled DNA through a series of orchestrated steps, DNA binding, cleavage, strand passage and religation. The N-terminal domain (NTD) of the type IA topoisomerases harbor DNA cleavage and religation activities, but the carboxyl terminal domain (CTD) is highly diverse. Most of these enzymes contain a varied number of Zn(2+) finger motifs in the CTD. The Zn(2+) finger motifs were found to be essential in Escherichia coli topoI but dispensable in the Thermotoga maritima enzyme. Although, the CTD of mycobacterial topoI lacks Zn(2+) fingers, it is indispensable for the DNA relaxation activity of the enzyme. The divergent CTD harbors three stretches of basic amino acids needed for the strand passage step of the reaction as demonstrated by a new assay. We also show that the basic amino acids constitute an independent DNA-binding site apart from the NTD and assist the simultaneous binding of two molecules of DNA to the enzyme, as required during the catalytic step. Although the NTD binds to DNA in a site-specific fashion to carry out DNA cleavage and religation, the basic residues in CTD bind to non-scissile DNA in a sequence-independent manner to promote the crucial strand passage step during DNA relaxation. The loss of Zn(2+) fingers from the mycobacterial topoI could be associated with Zn(2+) export and homeostasis.

| S-EPMC3753633 | biostudies-literature

Crystal structure of reverse gyrase: insights into the positive supercoiling of DNA.

Project description:Reverse gyrase is the only topoisomerase known to positively supercoil DNA. The protein appears to be unique to hyperthermophiles, where its activity is believed to protect the genome from denaturation. The 120 kDa enzyme is the only member of the type I topoisomerase family that requires ATP, which is bound and hydrolysed by a helicase-like domain. We have determined the crystal structure of reverse gyrase from Archaeoglobus fulgidus in the presence and absence of nucleotide cofactor. The structure provides the first view of an intact supercoiling enzyme, explains mechanistic differences from other type I topoisomerases and suggests a model for how the two domains of the protein cooperate to positively supercoil DNA. Coordinates have been deposited in the Protein Data Bank under accession codes 1GKU and 1GL9.

| S-EPMC125824 | biostudies-literature

Reverse gyrase--recent advances and current mechanistic understanding of positive DNA supercoiling.

Project description:Reverse gyrases are topoisomerases that introduce positive supercoils into DNA in an ATP-dependent reaction. They consist of a helicase domain and a topoisomerase domain that closely cooperate in catalysis. The mechanism of the functional cooperation of these domains has remained elusive. Recent studies have shown that the helicase domain is a nucleotide-regulated conformational switch that alternates between an open conformation with a low affinity for double-stranded DNA, and a closed state with a high double-stranded DNA affinity. The conformational cycle leads to transient separation of DNA duplexes by the helicase domain. Reverse gyrase-specific insertions in the helicase module are involved in binding to single-stranded DNA regions, DNA unwinding and supercoiling. Biochemical and structural data suggest that DNA processing by reverse gyrase is not based on sequential action of the helicase and topoisomerase domains, but rather the result of an intricate cooperation of both domains at all stages of the reaction. This review summarizes the recent advances of our understanding of the reverse gyrase mechanism. We put forward and discuss a refined, yet simple model in which reverse gyrase directs strand passage toward increasing linking numbers and positive supercoiling by controlling the conformation of a bound DNA bubble.

| S-EPMC4117796 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data