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Title: Electrostatics of DNA-DNA Juxtapositions: Consequences for Type II Topoisomerase Function


An abstract for this journal article is not available at this time.

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Research Org.:
Pacific Northwest National Laboratory (PNNL), Richland, WA (US), Environmental Molecular Sciences Laboratory (EMSL)
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Resource Type:
Journal Article
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Journal Name: Journal of Physics: Conference Series, 18(14):S173-S185
Country of Publication:
United States
Environmental Molecular Sciences Laboratory

Citation Formats

Randall, Graham L., Pettitt, Bernard M., Buck, Gregory R., and Zechiedrich, E. L. Electrostatics of DNA-DNA Juxtapositions: Consequences for Type II Topoisomerase Function. United States: N. p., 2006. Web. doi:10.1088/0953-8984/18/14/S03.
Randall, Graham L., Pettitt, Bernard M., Buck, Gregory R., & Zechiedrich, E. L. Electrostatics of DNA-DNA Juxtapositions: Consequences for Type II Topoisomerase Function. United States. doi:10.1088/0953-8984/18/14/S03.
Randall, Graham L., Pettitt, Bernard M., Buck, Gregory R., and Zechiedrich, E. L. Sun . "Electrostatics of DNA-DNA Juxtapositions: Consequences for Type II Topoisomerase Function". United States. doi:10.1088/0953-8984/18/14/S03.
title = {Electrostatics of DNA-DNA Juxtapositions: Consequences for Type II Topoisomerase Function},
author = {Randall, Graham L. and Pettitt, Bernard M. and Buck, Gregory R. and Zechiedrich, E. L.},
abstractNote = {An abstract for this journal article is not available at this time.},
doi = {10.1088/0953-8984/18/14/S03},
journal = {Journal of Physics: Conference Series, 18(14):S173-S185},
number = ,
volume = ,
place = {United States},
year = {Sun Mar 12 00:00:00 EST 2006},
month = {Sun Mar 12 00:00:00 EST 2006}
  • No abstract prepared.
  • Escherichia coli DNA topoisomerase III belongs to the type IA family of DNA topoisomerases, which transiently cleave single-stranded DNA (ssDNA) via a 5{prime} phosphotyrosine intermediate. We have solved crystal structures of wild-type E. coli topoisomerase III bound to an eight-base ssDNA molecule in three different pH environments. The structures reveal the enzyme in three distinct conformational states while bound to DNA. One conformation resembles the one observed previously with a DNA-bound, catalytically inactive mutant of topoisomerase III where DNA binding realigns catalytic residues to form a functional active site. Another conformation represents a novel intermediate in which DNA is boundmore » along the ssDNA-binding groove but does not enter the active site, which remains in a catalytically inactive, closed state. A third conformation shows an intermediate state where the enzyme is still in a closed state, but the ssDNA is starting to invade the active site. For the first time, the active site region in the presence of both the catalytic tyrosine and ssDNA substrate is revealed for a type IA DNA topoisomerase, although there is no evidence of ssDNA cleavage. Comparative analysis of the various conformational states suggests a sequence of domain movements undertaken by the enzyme upon substrate binding.« less
  • HL-60/AMSA is a human leukemia cell line that is 50-100-fold more resistant than its drug-sensitive HL-60 parent line to the cytotoxic actions of the DNA intercalator amsacrine (m-AMSA). HL-60/AMSA topoisomerase II is also resistant to the inhibitory actions of m-AMSA. HL-60/AMSA cells and topoisomerase II are cross-resistant to anthracycline and ellipticine intercalators but relatively sensitive to the nonintercalating topoisomerase II reactive epipodophyllotoxin etoposide. The authors now demonstrate that HL-60/AMSA and its topoisomerase II are cross-resistant to the DNA intercalators mitoxantrone and amonafide, thus strongly indicating that HL-60/AMSA and its topoisomerase II are resistant to topoisomerase II reactive intercalators but notmore » to nonintercalators. At high concentrations, mitoxantrone and amonafide were also found to inhibit their own, m-AMSA's, and etoposide's abilities to stabilize topoisomerase II-DNA complexes. These results suggest that the cytotoxicity of m-AMSA and etoposide is initiated primarily by the stabilization of the topoisomerase II-DNA complex. Other topoisomerase II reactive drugs may inhibit the enzyme at other steps in the topoisomerization cycle, particularly at elevated concentrations. Under these conditions, these latter drugs may not produce protein-associated DNA cleavage while still inhibiting topoisomerase II function as well as the actions of other topoisomerase II reactive drugs.« less
  • Mitoxantrone-resistant variants of the human HL-60 leukemia cell line are cross-resistant to several natural product and synthetic antineoplastic agents. The resistant cells (HL-60/MX2) retain sensitivity to the Vinca alkaloids vincristine and vinblastine, drugs that are typically associated with the classical multidrug resistance phenotype. Mitoxantrone accumulation and retention are equivalent in the sensitive and resistant cell types, suggesting that mitoxantrone resistance inn HL-60/MX2 cells might be associated with an alteration in the type II DNA topoisomerases. The authors discovered that topoisomerase II catalytic activity in 1.0 M NaCl nuclear extracts from the HL-60/MX2 variant was reduced 4- to 5-fold compared tomore » that in the parental HL-60 cells. Studies were designed to minimize the proteolytic degradation of the topoisomerase II enzymes by extraction of whole cells with hot SDS. When nuclear extracts from the two cell types were normalized for equivalent catalytic activity, mitoxantrone inhibited the decatenation of kDNA by these extracts to an equal extent but levels of mitoxantrone-induced cleavage of {sup 32}P-labeled pBR322 DNA by nuclear extracts from HL-60/MX2 cells were 3- to 4-fold lower than in comparable HL-60 extracts. Resistance to the topoisomerase II inhibitor mitoxantrone in HL-60/MX2 is associated with reduced nuclear and whole cell topoisomerase II catalytic activity, immunologically undetectable levels of the 180-kDa topoisomerase II isozyme, and reduced mitoxantrone-induced cleavage of radiolabeled DNA by topoisomerase II in nuclear extracts from these cells.« less
  • The structure of replicating simian virus 40 minichromosomes, extracted from camptothecin-treated infected cells, was investigated by biochemical and electron microscopic methods. The authors found that camptothecin frequently induced breaks at replication forks close to the replicative growth points. Replication branches were disrupted at about equal frequencies at the leading and the lagging strand sides of the fork. Since camptothecin is known to be a specific inhibitor of type I DNA topoisomerase, the authors suggest that this enzyme is acting very near the replication forks. This conclusion was supported by experiments with aphidicolin, a drug that blocks replicative fork movement, butmore » did not prevent the camptothecin-induced breakage of replication forks. The drug teniposide, and inhibitor of type II DNA topoisomerase, had only minor effects on the structure of these replicative intermediates.« less