Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Tuberculosis (TB) is one of the most important infections worldwide, with an estimated incidence of 10 million active cases per year. Rifampicin is a key component of the first-line therapy used in the treatment of tuberculosis. In Escherichia coli and Mycobacterium leprae, rifampicin has been shown to inhibit the beta subunit of RNA polymerase. The gene (rpoB) encoding this enzyme has been described in both species. We report the isolation of the homologous functional rifampicin resistance gene from M. tuberculosis. A library was constructed with 15 to 25 kb BamHI-digested DNA fragments from a rifampicin-resistant M. tuberculosis clinical isolate that was ligated into an E. coli-mycobacterial shuttle plasmid. Southern analysis of BamHI-digested DNA from 200 recombinant plasmids was performed and filters were hybridized to a 411 bp fragment of the beta subunit of RNA polymerase from M. tuberculosis. Only DNA from one plasmid (#86) hybridized, which suggested that the gene is found as a single copy per genome. This plasmid was able to transfer rifampicin resistance to sensitive M. smegmatis and thus codes for a functional genetic unit. Sequence analysis in the expected "hotspot" region in eight rifampicin-resistant M. tuberculosis strains (including one multidrug-resistant strain) revealed two novel mutations as well as others previously described.
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PMID:Isolation of the gene for the beta subunit of RNA polymerase from rifampicin-resistant Mycobacterium tuberculosis and identification of new mutations. 794 93

The interactions of transcription inhibitors with the open complex composed of Escherichia coli RNA polymerase and the lacUV5 promoter have been studied using gel retardation, the chemical nuclease activity of the cuprous complexes of 1,10-phenanthroline (OP) and its derivatives, and steady-state kinetics. Gel retardation shows that two inhibitors, the 2:1 2,9-dimethyl-1,10-phenanthroline-cuprous complex [(2,9-Me2OP)2Cu+] and rifampicin, bind stably to the open-complex. (2,9-Me2OP)2Cu+ blocks scission by the chemical nuclease by interfering with the binding of its redox-active isosteres. Rifampicin does not block scission by the cuprous complexes of 3,4,7,8-tetramethyl-OP, 4-phenyl-OP, and OP but does perturb scission by the cuprous complex of 5-phenyl-OP. Organic ligands including intercalating agents and groove binders (e.g., daunomycin, di(amidinophenyl)indole (DAPI), actinomycin D, distamycin, 9-aminoacridine, mithramycin, and chromomycin A3), which bind to free DNA with high affinity, do not form stable ternary complexes with the open-complex. Gel retardation experiments demonstrate that they promote dissociation of the enzyme from the promoter. The greater sensitivity of enzymatic catalysis to inhibitor concentration relative to polymerase binding suggests that these ligands form metastable, catalytically inactive ternary complexes with RNA polymerase and the promoter.
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PMID:Interactions of transcription inhibitors with the Escherichia coli RNA polymerase-lacUV5 promoter open complex. 811 83

The antimalarial activity of rifampicin, a specific inhibitor of bacterial ribonucleic acid (RNA) polymerase, was confirmed with Plasmodium falciparum in vitro and with P. chabaudi in vivo. The viability of ring forms of P. falciparum, measured by [3H]hypoxanthine and [14C]isoleucine uptake, was significantly reduced within 5 h of exposure to 2.5 microM rifampicin, the 50% inhibitory concentration. Streptolydigin and tagetitoxin, other specific inhibitors of bacterial RNA polymerase, were much less effective as antimalarials. A rifampicin-tolerant sub-line of P. falciparum was selected in vitro. When released from drug pressure, the tolerant line showed appreciably greater rates of incorporation of precursors and growth than the parent line, but over a period of months these characteristics gradually reverted. Rifampicin was effective against a chloroquine-resistant line of P. falciparum and the rifampicin-tolerant line had increased chloroquine sensitivity. Treatment of patent parasitaemias of P. chabaudi in mice with more than 100 mg/kg rifampicin twice daily significantly reduced the parasitaemia within 24 h and parasites were barely detectable on blood films by the fourth day. Recrudescence occurred on release of drug pressure.
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PMID:Antimalarial activity of rifampicin in vitro and in rodent models. 833 32

Rifampin is currently the most potent drug used in leprosy control programs. We show that the rifampin resistance which emerged in nine patients with lepromatous leprosy, who had received rifampin monotherapy, stemmed from mutations in the rpoB gene, which encodes the beta subunit of RNA polymerase of Mycobacterium leprae. In eight cases missense mutations were found to affect a serine residue, Ser-425, while in the remaining mutant a small insertion was found close to this site. These findings will be of use for the development of a rapid screening procedure, involving the polymerase chain reaction, for monitoring the emergence of rifampin-resistant M. leprae strains.
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PMID:Molecular basis of rifampin resistance in Mycobacterium leprae. 846 Sep 11

Rifampicin-resistant (Rifr) mutations of Escherichia coli map to the central portion of the rpoB gene, which encodes the beta subunit of RNA polymerase. These mutations are located in three distinct clusters, designated I, II and III. Three intragenic suppressors of the cluster III Rifr mutation, rpoB3406(RH687), restore the ability of the mutant strain to grow at low and high temperatures and map to a single locus in cluster I. These suppressors are identical to two previously characterized Rifr alleles, rpoB3401(RC529) and rpoB3402(RS529). None of the other 14 previously identified Rifr mutations that we have characterized confers this phenotype. We suggest that this allele-specific suppression results from interaction between Cluster I and Cluster III of the beta subunit.
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PMID:Genetic evidence for the interaction between cluster I and cluster III rifampicin resistant mutations. 849 55

Rifampin resistance in respiratory isolates of Mycobacterium tuberculosis from Mozambique was detected by screening for point mutations using polymerase chain reaction (PCR) and DNA sequence analysis. The target template was a 350-bp fragment of rpoB encoding the beta-subunit of the RNA polymerase. Of the 66 strains studied, 38 were rifampin resistant by susceptibility testing with the radiometric method, 3 were intermediately resistant, and 25 were susceptible to rifampin. In 39 of the 41 rifampin-resistant strains, base-substitutions in the rpoB fragment were detected, and a total of 13 distinct mutations affecting 6 amino acids were observed. One of these mutations (His-->Thr in amino acid 526) was not previously described. The isolates were also investigated by restriction fragment length polymorphism (RFLP) analysis using the insertion element IS6110 as a hybridization probe. A total of 47 RFLP patterns were identified, with up to 9 isolates having the same RFLP pattern. Strains with the same RFLP pattern harbored different mutations in rpoB, suggesting that acquisition of rifampin resistance followed the spread of a rifampin-susceptible clone. The data showed that rifampin resistance can be detected with a high sensitivity by DNA sequence analysis of this fragment of rpoB. However, a few strains with rifampin resistance due to factors other than base substitutions in rpoB could be missed.
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PMID:Detection of rifampin resistance among isolates of Mycobacterium tuberculosis from Mozambique. 915 3

Rifampicin resistance has arisen in several different species of bacteria because of alterations to one or more regions in the target of the antibiotic, the beta-subunit of RNA polymerase encoded by rpoB. Nucleotide sequence analysis of a 270 bp fragment of rpoB from 16 clinical rifampicin-susceptible isolates of Streptococcus pneumoniae, 8 clinical rifampicin-resistant isolates, and 3 spontaneous rifampicin-resistant mutants, has revealed that, as with previously examined species, point mutations within the cluster I region of rpoB, at sites encoding Asp516 and HiS526, also confer resistance to rifampicin in this important human pathogen. Moreover, the residues within cluster I, that were altered within the rifampicin-resistant mutants of S. pneumoniae, were in the same position as those previously found to alter in resistant isolates of Escherichia coli and Mycobacterium tuberculosis. Sequence analysis of rpoB, both from these isolates of S. pneumoniae and from two strains of S. mitis, reveals that, among a number of clinical isolates, resistance to rifampicin in S. pneumoniae has arisen by point mutation. However, the nucleotide sequence of rpoB from one isolate examined suggests that interspecies gene transfer may also have played a role in the evolution of rifampicin-resistance in S. pneumoniae.
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PMID:Molecular evolution of rifampicin resistance in Streptococcus pneumoniae. 953 28

A centrifugation method was used to investigate the accumulation of 14C-rifampicin by Staphylococcus aureus and Escherichia coli, and to characterize the mechanism of rifampicin transport into S. aureus. For both species, drug accumulation was rapid with the steady-state concentration (SSC) reached within 40 s of drug exposure. Rifampicin accumulation by S. aureus was temperature and pH dependent; the lower the experimental temperature and the lower the experimental pH, the lower was the concentration of rifampicin accumulated. Accumulation was unaffected by the presence of inhibitors of antibiotic efflux, carbonyl cyanide m-chlorophenylhydrazone (CCCP), dinitrophenol (DNP), or reserpine. Exposure to increasing concentrations of rifampicin suggested that the accumulation process was saturable above a rifampicin concentration of 0.2 mg/L. Michaelis-Menten kinetics gave an apparent Km and Vmax for rifampicin, calculated from a Lineweaver-Burk plot, of 0.05 mg/L (0.06 microM) and 3.8 ng rifampicin per second, respectively. However, calculations suggest that these values reflect those for binding of rifampicin to its target, RNA polymerase.
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PMID:Accumulation of rifampicin by Escherichia coli and Staphylococcus aureus. 984 43

Rifampicin is an antibiotic which binds to the beta subunit of prokaryotic RNA polymerases and prevents initiation of transcription. It was found previously that production of heat shock proteins in Escherichia coli cells after a shift from 30 degrees C to 43 degrees C is not completely inhibited by this antibiotic. Here we demonstrate that while activity of a pL-lacZ fusion (pL is a sigma70-dependent promoter) in E. coli cells is strongly inhibited by rifampicin, a p(groE)-lacZ fusion, whose activity is dependent on the sigam32 factor, retains significant residual activity even at relatively high rifampicin concentrations. Differential sensitivity to this antibiotic of RNA polymerase holoenzymes containing either the sigma70 or the sigma32 subunit was confirmed in vitro. Since the effects of an antibiotic that binds to the beta subunit can be modulated by the presence of either the sigma70 or the sigma32 subunit in the holoenzyme, it is tempting to speculate that binding of various sigma factors to the core of RNA polymerase results in different conformations of particular holoenzymes, including changes in the core enzyme.
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PMID:Differential inhibition of transcription from sigma70- and sigma32-dependent promoters by rifampicin. 986 49

The sequence of the rpoB gene from Listeria monocytogenes was determined. Rifampin-resistant (Rif(r)) mutants arising from L. monocytogenes cultures exposed to rifampin were isolated, and by partial sequencing of their rpoB genes, seven different point mutations affecting five different amino acids (473Asp-->Asn or Gly, 479Gly-->Asp, 483His-->Tyr or Leu, 528Ile-->Phe, and 530Ser-->Tyr), which led to MICs of 0.5 to 100 microg/ml for the organisms, were determined. These mutants showed various deficiencies for growth at 42 degrees C, with only one being comparable to the wild-type strain. The interaction of these Rif(r) mutants with human Caco-2 cells was examined by using an immunofluorescence technique. Three mutants failed to interact, while three showed a reduced interaction compared to that of the wild type. It is believed that these pleiotropic phenotypes have arisen as a result of mutations within the DNA-dependent RNA polymerase holoenzyme.
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PMID:Isolation of rifampin-resistant mutants of Listeria monocytogenes and their characterization by rpoB gene sequencing, temperature sensitivity for growth, and interaction with an epithelial cell line. 1044 75


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