The MukB-ParC Interaction Affects the Intramolecular, Not Intermolecular, Activities of Topoisomerase IV

Hayama, Ryo; Bahng, Soon; Karasu, Mehmet E.; Marians, Kenneth J.

doi:10.1074/jbc.m112.418087

Cited by 36 publications

(58 citation statements)

References 40 publications

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“…We had previously engineered and purified a MukB variant that was defective in DNA binding (MukB dna (16), R187E/ R189E). These two amino acid substitutions are the equivalent of the R216E and R218E mutations made in Haemophilus ducreyi MukB that elicited DNA binding deficiency (13).…”

Section: Resultsmentioning

confidence: 99%

“…We have reported (16) that this interaction stimulates the intramolecular activities of Topo IV, negative supercoil relaxation and knotting, but not the intermolecular activities of Topo IV, catenation/decatenation of DNA rings; whereas Berger, Oakley and colleagues (15) have shown a moderate stimulation by MukB of Topo IV-catalyzed decatenation of Crithidia mitochondrial DNA.…”

mentioning

confidence: 76%

“…DNAs and Proteins-MukB and MukB dna (MukB R187E/ R189E) were prepared as described (14,16). Topo III was prepared as described (35).…”

Section: Methodsmentioning

confidence: 99%

“…Plasmid pCG09 is 11,088 bp long and was described in Ref. 16. Gapped pCG09 DNA was prepared as follows.…”

Section: Methodsmentioning

confidence: 99%

“…We have proposed that the Topo IV-MukB interaction is necessary for efficient chromosome condensation, with Topo IV possibly stabilizing MukB interactions with distal regions of the chromosome to form loops in the DNA (16). Disruption of this interaction would thereby cause chromosome decondensation that could then disrupt proper chromosome decatenation.…”

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confidence: 99%

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MukB-mediated Catenation of DNA Is ATP and MukEF Independent

Bahng

Hayama

Marians

2016

Journal of Biological Chemistry

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Edited by Patrick SungProperly condensed chromosomes are necessary for accurate segregation of the sisters after DNA replication. The Escherichia coli condesin is MukB, a structural maintenance of chromosomes (SMC)-like protein, which forms a complex with MukE and the kleisin MukF. MukB is known to be able to mediate knotting of a DNA ring, an intramolecular reaction. In our investigations of how MukB condenses DNA we discovered that it can also mediate catenation of two DNA rings, an intermolecular reaction. This activity of MukB requires DNA binding by the head domains of the protein but does not require either ATP or its partner proteins MukE or MukF. The ability of MukB to mediate DNA catenation underscores its potential for bringing distal regions of a chromosome together.The Escherichia coli chromosome is condensed about 1000-fold to fit into the nucleoid of the bacterium. A number of factors contribute to this extreme DNA condensation, among them are: DNA supercoiling, the binding of various nucleoid associated proteins like HU, Fis, and H-NS, and the binding of the structural maintenance of chromosomes (SMC) 3 -like condensin MukBEF (1, 2). SMC proteins act to manage the shape and behavior of chromosomes in both prokaryotes and eukaryotes (3). These proteins dimerize at a hinge region that is flanked by long coiled-coil regions that can be 40 -50 nm in length and that end in head domains that bind and hydrolyze ATP, as well as the bridging kleisin protein (MukF for the E. coli condensin (4)). The kleisin is then itself bound by another protein (MukE for the E. coli condensin (4)). There are three versions of the eukaryotic SMC proteins: the condensin, required for packaging of the chromosomes and comprised of SMC2 and SMC4; the cohesin, required for holding sister chromosomes together during mitosis and comprised of SMC1 and SMC3; and the SMC5-SMC6 complex, required for various aspects of DNA repair (3).It is generally acknowledged that the eukaryotic condensin and cohesin trap DNA topologically in the protein triangle formed by the SMC proteins and the kleisin (5, 6), although an alternative model for DNA binding for the eukaryotic cohesin exists (7). Recent reports suggest that the Bacillus subtilis SMC protein (8) and MukB (9) also trap chromosomes topologically. MukB binds linear and circular DNA in vitro and can induce negative supercoils and knots in relaxed circular DNA in the presence of a topoisomerase (10). Binding of MukB to chromosomal DNA in vivo requires ATP and MukEF (11, 12), although MukB DNA binding in vitro does not (10, 13).We (14) and the Burger and Oakley (15) labs have shown that the MukB hinge region interacts with the C-terminal ␤-propeller region of the ParC subunit of the cellular decatenase topoisomerase IV (Topo IV). We have reported (16) that this interaction stimulates the intramolecular activities of Topo IV, negative supercoil relaxation and knotting, but not the intermolecular activities of Topo IV, catenation/decatenation of DNA rings; whereas Berger, Oakley and colleag...

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 76%

“…DNAs and Proteins-MukB and MukB dna (MukB R187E/ R189E) were prepared as described (14,16). Topo III was prepared as described (35).…”

Section: Methodsmentioning

confidence: 99%

“…Plasmid pCG09 is 11,088 bp long and was described in Ref. 16. Gapped pCG09 DNA was prepared as follows.…”

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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MukB-mediated Catenation of DNA Is ATP and MukEF Independent

Bahng

Hayama

Marians

2016

Journal of Biological Chemistry

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References

2020

Structure and Function of the Bacterial Genome

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The Localization and Action of Topoisomerase IV in Escherichia coli Chromosome Segregation Is Coordinated by the SMC Complex, MukBEF

et al. 2015

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SummaryThe type II topoisomerase TopoIV, which has an essential role in Escherichia coli chromosome decatenation, interacts with MukBEF, an SMC (structural maintenance of chromosomes) complex that acts in chromosome segregation. We have characterized the intracellular dynamics of individual TopoIV molecules and the consequences of their interaction with MukBEF clusters by using photoactivated-localization microscopy. We show that ∼15 TopoIV molecules per cell are associated with MukBEF clusters that are preferentially localized to the replication origin region (ori), close to the long axis of the cell. A replication-dependent increase in the fraction of immobile molecules, together with a proposed catalytic cycle of ∼1.8 s, is consistent with the majority of active TopoIV molecules catalyzing decatenation, with a minority maintaining steady-state DNA supercoiling. Finally, we show that the MukB-ParC interaction is crucial for timely decatenation and segregation of newly replicated ori DNA.

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The MukB-ParC Interaction Affects the Intramolecular, Not Intermolecular, Activities of Topoisomerase IV

Cited by 36 publications

References 40 publications

MukB-mediated Catenation of DNA Is ATP and MukEF Independent

MukB-mediated Catenation of DNA Is ATP and MukEF Independent

References

The Localization and Action of Topoisomerase IV in Escherichia coli Chromosome Segregation Is Coordinated by the SMC Complex, MukBEF

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