Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The solution structure of Escherichia coli DNA gyrase, an enzyme that catalyzes the ATP-dependent supercoiling of DNA, has been characterized by small-angle neutron scattering (SANS) and dynamic light-scattering (DLS). The enzyme and its complex with a 172 base-pair fragment of duplex DNA, in H2O or 2H2O solvent, were studied by contrast variation and the measurement of hydrodynamic parameters as a function of scattering angle. The complex was also measured in the presence of 5'-adenylyl-beta,gamma-imidodiphosphate (ADPNP), a non-hydrolyzable ATP analog that is known to support limited supercoiling. The values of the radius of gyration, Rg = 67 A, from SANS and the hydrodynamic radius, Rh = 64 A, from DLS predict a larger than expected volume for the enzyme, supporting the notion of channels or cavities within the molecule. In addition, several classes of models were rejected based on SANS data obtained in 2H2O at larger scattering angles. The best fit to both the SANS and DLS data is obtained for oblate, inhomogeneous particles approximately 175 A wide and 52 A thick. Such particles provide a large surface area for DNA interaction. Both Rg and Rh values change very little upon addition of DNA, suggesting that DNA binds in a manner that does not significantly change the shape of the protein. No appreciable change in structure is found with the addition of ADPNP. However, the higher-angle SANS data indicate a slight rearrangement of the enzyme in the presence of nucleotide.
J Mol Biol 1990 Jan 05
PMID:Neutron and light-scattering studies of DNA gyrase and its complex with DNA. 215 34

A multidrug-resistant variant of the P388 leukemia cell line exhibits multiple biochemical changes, including reduced drug accumulation and markedly reduced DNA strand breakage induced by anthracyclines. To investigate whether the reduced formation of drug-induced DNA breaks was due to alteration of DNA topoisomerase II activity, nuclear extracts and partially purified enzymes from the sensitive line and the resistant subline were compared. DNA topoisomerase II activity in 0.35 M NaCl nuclear extracts from sensitive cells was approximately 1.7 times higher than that found in extracts from resistant cells, as determined by ability to unknot P4 phage DNA. In addition, it was found that teniposide-stimulated topoisomerase II DNA cleavage activity of nuclear extract from resistant cells was at least 10-fold lower than that from sensitive cells. This differential sensitivity paralleled a similar drug response of nuclei, as determined by the alkaline elution method. However, partially purified DNA topoisomerase II showed similar drug sensitivity in both cell lines. This finding suggests the presence of a modulating factor, which may be lost during purification. These results, indicating a reduction of both catalytic activity and DNA cleavage activity of DNA topoisomerase II in P388 multidrug-resistant cells, emphasize the importance of DNa topoisomerase function in the resistance mechanism. Thus, the concomitant involvement of multiple mechanisms could explain the high degree of resistance of these cells.
Mol Pharmacol 1990 Jan
PMID:Evidence of DNA topoisomerase II-dependent mechanisms of multidrug resistance in P388 leukemia cells. 215 5

A mixed oligonucleotide probe containing sequences encoding a septapeptide found in yeast, Drosophila and human DNA topoisomerase II was used to screen a genomic library of Trypanosoma brucei. A positive was obtained, and nucleotide sequencing shows that the entire gene encoding DNA topoisomerase II of this organism, TbrTOP2, resides within the T. brucei insert of the clone. A single open reading frame of 1221 triplet codons starting from the first ATG was identified; the amino acid sequence deduced from it is highly homologous to other eukaryotic DNA topoisomerase II and corresponds to a 137-kDa polypeptide. Analysis of restriction endonuclease digests of T. brucei DNA by blot hybridization following gel electrophoresis indicates that TbrTOP2 is a single-copy gene.
Mol Biochem Parasitol 1990 Jan 01
PMID:The TOP2 gene of Trypanosoma brucei: a single-copy gene that shares extensive homology with other TOP2 genes encoding eukaryotic DNA topoisomerase II. 215 53

A system for detecting a spontaneous deletion in Escherichia coli was developed and the role of DNA gyrase in deletion formation was studied. A derivative of lambda plac5, lambda AM36, was isolated in which whole pBR322 DNA was inserted in the lacZ gene and 227 bp of the lac gene duplicated at both sides of the pBR322 DNA. E. coli lac- strains lysogenized by lambda AM36 had a Lac- phenotype and segregated Lac+ revertants. Sequence analyses showed that the revertant was formed by a deletion that eliminated the inserted pBR322 DNA and one copy of the duplicated segments. The frequency of lac+ revertant formation was independent of recA function, was increased by oxolinic acid, an inhibitor of DNA gyrase, but was not increased in a lysogen of a nalidixic acid-resistant derivative. The reversion frequencies of temperature sensitive mutants of gyrA gene are 10 to 100 times lower than that of the wild-type strain. These results indicate that the DNA gyrase of E. coli participated in the in vivo deletion formation resulting from the direct repeats.
Mol Gen Genet 1990 Feb
PMID:The DNA gyrase of Escherichia coli participates in the formation of a spontaneous deletion by recA-independent recombination in vivo. 216 49

The effect of cyanomorpholinyldoxorubicin, morpholinyldoxorubicin, doxorubicin, and Actinomycin D were studied on purified mouse leukemia (L1210) DNA topoisomerases I and II. DNA unwinding and cross-linking were also studied. It was found that 1) morpholinyldoxorubicin, cyanomorpholinyldoxorubicin, and Actinomycin D (but not doxorubicin) stimulated DNA topoisomerase I-induced cleavage at specific DNA sites; 2) only doxorubicin and Actinomycin D stimulated DNA cleavage by DNA topoisomerase II; 3) at higher drug concentrations, DNA intercalators suppressed enzyme-mediated DNA cleavage induced by DNA topoisomerase I, as well as topoisomerase II; 4) only cyanomorpholinyldoxorubicin produced DNA-DNA cross-links; no DNA unwinding could be observed; and 5) DNA intercalation (unwinding) potency of morpholinyldoxorubicin was about 2-fold less than that of doxorubicin. The data indicate that some DNA intercalators are not only inhibitors of DNA topoisomerase II but act also on DNA topoisomerase I. The stabilization of cleavage intermediates by intercalators may have a common mechanism for DNA topoisomerase I and DNA topoisomerase II.
Mol Pharmacol 1990 Jul
PMID:Effects of morpholinyl doxorubicins, doxorubicin, and actinomycin D on mammalian DNA topoisomerases I and II. 216 30

Stimulation of gyrA expression in an in vitro transcription-translation system by novobiocin, a DNA gyrase inhibitor, depends on the DNA concentration in the extract. At low DNA concentrations (less than 20 micrograms/ml) we note significant stimulation (2 to 25 x) upon the addition of novobiocin; at high DNA concentrations, stimulation is minimal. This observation is not due to the limited capacity of the system to transcribe or relax DNA. Using an extract prepared from a novobiocin-resistant strain of Escherichia coli, we were able to show that DNA gyrase mediates the novobiocin-enhanced expression. In an experiment with a fixed level of a gyrA-lac template, we found that the addition of a second non-Lac template increased expression from the gyrA-lac template, while concomitantly decreasing the extent of novobiocin stimulation. These observations are consistent with an inhibitory factor that can be titrated by increasing the DNA concentration and whose effects are minimized when the DNA template is in a relaxed conformation. The results of a mixed-extract experiment using two extracts that differ in their activities and degrees of novobiocin stimulation are also consistent with an inhibitory factor that mediates the relaxation-induced stimulation of transcription.
J Mol Biol 1990 Jul 20
PMID:Inhibition of DNA gyrase activity in an in vitro transcription-translation system stimulates gyrA expression in a DNA concentration dependent manner. Evidence for the involvement of factors which may be titrated. 216 66

Mortality among cystic fibrosis (CF) patients is most commonly attributed to pulmonary infection by mucoid, alginate-producing Pseudomonas aeruginosa. The initial infecting P. aeruginosa are typically non-mucoid; however, upon continued exposure to the CF lung environment, they become highly mucoid. The CF lung is an osmotically high environment because of the presence of substantial concentrations of electrolytes and dehydrated mucus. In this report we demonstrate that ethanol (a commonly used dehydrating agent) activates transcription from a critical alginate promoter, algD, and show that prolonged exposure to ethanol allows switching to the mucoid form. This activation appears to be dependent on DNA gyrase. Analysis of alginate gene activation, and the subsequent reversal of the activation process by bacterial DNA gyrase inhibitors, should aid the development of treatment strategies for CF patients infected with this organism.
Mol Microbiol 1990 May
PMID:Pulmonary dehydration and infection in cystic fibrosis: evidence that ethanol activates alginate gene expression and induction of mucoidy in Pseudomonas aeruginosa. 216 23

Studies have suggested that recombinant tumor necrosis factor-alpha (TNF-alpha) may potentiate the killing of murine tumor cells by drugs targeted at DNA topoisomerase II. We have examined the combined cytotoxic effects of the topoisomerase-targeted drug etoposide and TNF in small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) cell lines using clonogenic assays and a novel flow cytometry technique relying on differential uptake of fluorescein diacetate (FDA) and propidium iodide (PI) by viable and nonviable cells. Good correlation of IC50 determinations for etoposide were noted between clonogenic assays and the FDA/PI technique for both classic and variant SCLC cell lines. The effects of etoposide on the classic SCLC line H209 were potentiated by TNF with a decrease in the IC50 from 3.3 microM to 1.0 microM as determined by FDA/PI. Tumor necrosis factor alone had little effect on the growth or cloning efficiency of H209 cells. Tumor necrosis factor alone stimulated the growth and cloning of variant SCLC line N417, but the cytotoxicity of etoposide was not potentiated by TNF in N417 cells. Tumor necrosis factor alone inhibited the growth and cloning of the NSCLC line H125 but exerted a marked protective effect against higher concentrations of etoposide. It appears that the interaction of TNF with etoposide varies between cell lines and between subclasses of human lung cancer.
Mol Biother 1990 Sep
PMID:Interaction of recombinant human tumor necrosis factor and etoposide in human lung cancer cell lines. 217 61

The 64 x 10(3) Mr N-terminal breakage-reunion domain of the Escherichia coli DNA gyrase A protein was purified from an over-expressing strain. When complexed with the gyrase B protein, this truncated A protein has all of the enzymic properties of the full-length counterpart, although with reduced efficiency in some cases. The 64 x 10(3) Mr protein has been crystallized in several forms, a number of which were too small for crystallographic analysis. However, two forms grew to sufficient size for preliminary X-ray analysis. Both forms were tetragonal with a primitive lattice. One form (type I) had cell dimensions of a = b = 170 A, c = 145 A a space group of either P41212 (P43212) or P42212, and diffracted to 6 A resolution. The type II crystals had cell dimensions of a = b = 177 A, c = 175 A, a space group of P41212 (P43212) or P42212, and diffracted to at least 4.5 A resolution. Both crystal forms apparently contained four subunits (possibly a tetramer) in the asymmetric unit. We are attempting to increase the size and quality of these crystals.
J Mol Biol 1990 Oct 20
PMID:Preliminary crystallographic analysis of the breakage-reunion domain of the Escherichia coli DNA gyrase A protein. 217 50

We have identified a clone from a lambda EMBL3 library containing a 19kb insert of Mycoplasma pneumoniae DNA which includes the genes that encode both subunits of DNA gyrase. The gyrB gene and the 5' end of the gyrA gene have been subcloned into M13. The gyrB gene is 1953bp in length and overlaps the gyrA gene by a single base. The nucleotide sequence of these subclones has significant homology to previously reported gyrase genes. In terms of the size of the gyrB gene and its proximity to the gyrA gene, M. pneumoniae is more similar to Bacillus subtilis than to Escherichia coli.
Mol Microbiol 1990 Jul
PMID:Mycoplasma pneumoniae DNA gyrase genes. 217 93


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