Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

Mycoplasma hominis, an opportunistic pathogenic bacterium of humans, has a small genome of 700 kb. Despite this, multiple copies of gene sequences with similarities to the structural gene (lmp1) of a 135-kDa surface-located membrane protein (Lmp1) have been identified on the genome of M. hominis PG21 (lmp2, lmp3, and lmp4). The distance between the lmp1-lmp2 region and the lmp3-lmp4 region was more than 110 kb. lmp3-lmp4 of M. hominis PG21 was sequenced and found to contain two putative genes. The gene region of 6.5 kb contained a 5' unique region and a 3' unique region separated by 9 0.5-kb repeats with 51 to 90% similarity to 10 similar repeats found in the lmp1-lmp2 region. The 0.5-kb DNA repeats thus comprised about 1% of the entire genome. In both regions, a base change in one of the repeats gave rise to a stop codon, and thereby lmp2 and lmp4 occurred. By PCR amplification of reverse-transcriptase-generated cDNA it was shown that all four genes were transcribed. By use of Lmp-specific antibodies we showed that both lmp1 and lmp3 were translated into proteins (Lmp1 and Lmp3). Each of the four lmp genes represented by their unique cloned segments was used as a probe to analyze the presence, distribution, and organization of the genes within the genome in 13 M. hominis isolates. The repetitive element was detected at one or two locations on the chromosome for all isolates. The lmp3-specific element was present in all isolates, and lmp1- and lmp2-specific elements were present in all but one isolate. The lmp4-specific element was present in about half the isolates tested. For five M. hominis isolates the chromosomal location of the lmp genes was mapped.
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PMID:Analysis of 0.5-kilobase-pair repeats in the Mycoplasma hominis lmp gene system and identification of gene products. 863 64

The past decade has seen the rapid advancement of molecular biology and its application in the field of infectious diseases. The polymerase chain reaction (PCR) is a technique which brings about the in vitro amplification of DNA, and is clinically useful in the sensitive, specific and rapid diagnosis of various infectious diseases. The fast diagnosis of viral infections using PCR is a prime example, since viral culture may take weeks to grow and since serologic conversion seldom occurs until the convalescent phase of the clinical illness. Similarly, PCR is applicable to the diagnosis of pulmonary infections caused by mycobacteria, Legionella, methicillin-resistant Staphylococcus aureus, anaerobes, Mycoplasma, Chlamydia, Pneumocystis carinii and so on. On the other hand, the drug-resistant phenotype of a microorganism is predictable by the detection of gene alterations related to the drug resistance. For example, the deletion of the catalase-peroxidase gene relating to isoniazid resistance of mycobacteria, and point mutations in the RNA polymerase beta subunit (rpoB) gene relating to rifampicin resistance have been elucidated. As these applied techniques become more widely available and less costly, they should contribute not only to the rapid diagnosis of infectious diseases, but also to the adequate selection of antibiotics and the shortening of hospitalization.
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PMID:[Advances in genetic diagnostics for respiratory tract infections]. 883 14

The Streptomyces coelicolor rpoC gene, that encodes the beta' subunit of RNA polymerase, was isolated using the Escherichia coli rpoC gene as a hybridization probe. Comparison of the predicted amino acid sequence of the S. coelicolor beta' subunit to those characterized from other bacteria revealed three distinct subfamilies of beta' subunits, one of which consists of the S. coelicolor subunit and those from Mycobacterium leprae and Mycoplasma genitalium. Using site-directed mutagenesis, the carboxy terminus of the S. coelicolor beta' subunit was modified to contain six histidine residues. The histidine-tagged gene, rpoCHIS, was used to replace the wild-type allele in the chromosome of S. coelicolor and S. lividans. These strains were unaffected in growth and sporulation, demonstrating that the histidine-tagged RNA polymerase was competent to carry out all essential in-vivo functions. During a 1-day procedure, highly purified RNA polymerase was obtained by nickel-NTA agarose affinity chromatography followed by heparin-sepharose chromatography. Using in-vitro run-off transcription assays, the affinity purified RNA polymerase was shown to initiate transcription correctly from the S. lividans galP1 and galP2 promoters, and the Bacillus subtilus veg and ctc promoters. An extension of this procedure yielded highly-purified core RNA polymerase. To facilitate introduction of the rpoCHIS allele into other genetic backgrounds, a mutation in the adjacent gene, rpoB (rifA), conferring rifampin-resistance, was isolated in S. coelicolor to provide a genetic marker to follow transfer of the rpoCHIS allele. The use of this affinity chromatography procedure, in combination with the ability to introduce the rpoCHIS allele into different Streptomyces strains by transformation, will greatly facilitate the in-vitro analysis of transcription in members of this genus.
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PMID:Characterization of the rpoC gene of Streptomyces coelicolor A3(2) and its use to develop a simple and rapid method for the purification of RNA polymerase. 932 38

The phylogenetic placement of the Aquifex and Thermotoga lineages has been inferred from (i) the concatenated ribosomal proteins S10, L3, L4, L23, L2, S19, L22, and S3 encoded in the S10 operon (833 aa positions); (ii) the joint sequences of the elongation factors Tu(1alpha) and G(2) coded by the str operon tuf and fus genes (733 aa positions); and (iii) the joint RNA polymerase beta- and beta'-type subunits encoded in the rpoBC operon (1130 aa positions). Phylogenies of r-protein and EF sequences support with moderate (r-proteins) to high statistical confidence (EFs) the placement of the two hyperthermophiles at the base of the bacterial clade in agreement with phylogenies of rRNA sequences. In the more robust EF-based phylogenies, the branching of Aquifex and Thermotoga below the successive bacterial lineages is given at bootstrap proportions of 82% (maximum likelihood; ML) and 85% (maximum parsimony; MP), in contrast to the trees inferred from the separate EF-Tu(1alpha) and EF-G(2) data sets, which lack both resolution and statistical robustness. In the EF analysis MP outperforms ML in discriminating (at the 0.05 level) trees having A. pyrophilus and T. maritima as the most basal lineages from competing alternatives that have (i) mesophiles, or the Thermus genus, as the deepest bacterial radiation and (ii) a monophyletic A. pyrophilus-T. maritima cluster situated at the base of the bacterial clade. RNAP-based phylogenies are equivocal with respect to the Aquifex and Thermotoga placements. The two hyperthermophiles fall basal to all other bacterial phyla when potential artifacts contributed by the compositionally biased and fast-evolving Mycoplasma genitalium and Mycoplasma pneumoniae sequences are eschewed. However, the branching order of the phyla is tenuously supported in ML trees inferred by the exhaustive search method and is unresolved in ML trees inferred by the quartet puzzling algorithm. A rooting of the RNA polymerase-subunit tree at the mycoplasma level seen in both the MP trees and the ML trees reconstructed with suboptimal amino acid substitution models is not supported by the EF-based phylogenies which robustly affiliate mycoplasmas with low-G+C gram-positives and, most probably, reflects a "long branch attraction" artifact.
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PMID:Phylogenetic depth of the bacterial genera Aquifex and Thermotoga inferred from analysis of ribosomal protein, elongation factor, and RNA polymerase subunit sequences. 1079 28

The aim of this study was to generate urgently needed data on respiratory pathogens in German children using an economical and efficient tool. Nasopharyngeal aspirates of hospitalized children 0-16 years of age with an acute respiratory tract infection were tested by a nine-valent multiplex reverse-transcriptase polymerase chain reaction. Of 1281 children, 449 (35%) had an acute respiratory tract infection caused by at least one of the organisms studied; there were 29 cases of dual infection. At least 34-42% of severe acute respiratory tract infections in children under 5 years of age were caused by viruses. In children over 5 years of age, this proportion was 23% (P<0.001). Infection during the first 2 years of life was most frequently due to respiratory syncytial virus (n = 162 cases). Parainfluenza virus type 3 (n = 22) and type 1 (n = 14) were detected almost exclusively in children under 5 years of age. Influenza A (n = 90) and adenoviruses (n = 98) were prevalent in all age groups. The frequency of influenza B virus isolation (n = 17) rose significantly after the age of 5 years. Mycoplasma pneumoniae infection (n = 24 cases, 5.2%) was most frequent in 5- to 16-year-old patients. Only one case of Chlamydia pneumoniae infection was found. Since the distribution of pathogens within the different types of lower respiratory tract infections is very similar, it seems that host factors determine which form of lower respiratory tract infection develops in an individual patient. The multiplex reverse-transcriptase polymerase chain reaction may, in the future, become an important tool for epidemiological studies as well as for individual diagnosis.
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PMID:Epidemiological investigation of nine respiratory pathogens in hospitalized children in Germany using multiplex reverse-transcriptase polymerase chain reaction. 1089 33

Until recently no isolate of Tropheryma whippelii was available, and therefore genetic studies were limited to those based on PCR amplification of conserved genes. In this study we determined the nucleotide sequence of rpoB (encoding the beta-subunit of RNA polymerase) from a cultured strain of T. whippelii using degenerate consensus PCR and genome walking. The T. whippelii rpoB consists of 3,657 bp with a 50.4% GC content and encodes 1,218 amino acids with a calculated molecular mass of 138 kDa. Comparison of T. whippelii RpoB with other eubacterial RpoB proteins indicated sequence similarity ranging from 57.19 (Mycoplasma pneumoniae) to 74.63% (Mycobacterium tuberculosis). Phylogenetic analysis of T. whippelii based on comparison of its RpoB sequence with sequences available for other bacteria was consistent with that previously derived from the 16S ribosomal DNA (rDNA) sequence, indicating that it belongs to the actinomyces clade. The sequence comparison allowed the design of a primer pair, TwrpoB.F and TwrpoB.R, specific for T. whippelii rpoB. When incorporated into a PCR, this primer pair allowed the detection of T. whippelii rpoB in three of three 16S rDNA PCR-positive biopsy specimens and zero of seven negative controls. rpoB could therefore be targeted in PCR-mediated detection and identification of this emerging bacterial species. This approach has previously been shown useful for the identification of related mycobacteria. This study underscores that a method involving isolation and then propagation of emerging bacteria is a useful way to quickly achieve extensive molecular knowledge of these pathogens.
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PMID:rpoB sequence analysis of cultured Tropheryma whippelii. 1142 49

Two conservative gene clusters, the S10 ribosomal protein region and one (of the two) set of rRNA genes, were split in a genome crossover rearrangement event in Mycoplasma gallisepticum. As a result of the rearrangement the major part of the S10 ribosomal protein cluster is located upstream of genes for 23S-5S rRNA (rrn23-5), but the genes infA-rpl36-rps13-rpoA-rpl17 are located immediately downstream of the isolated gene for 16S rRNA (rrn16). A new ribosomal protein cluster infA-rpl36-rps13-rpoA-rpl17-rps16-trmD-rpl19 was formed. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed that this ribosomal protein cluster is an operon.
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PMID:Mycoplasma gallisepticum rpoA gene cluster. 1195 50

In a case-control study of the infectious agents associated with natural outbreaks of respiratory disease in pheasants, 28 batches of birds from sites affected by disease and eight batches of birds from unaffected sites were examined by six veterinary laboratories in England, Wales and Scotland, and tested for mycoplasmas, other bacteria and viruses. Sinusitis was the commonest sign of disease and was associated with Mycoplasma gallisepticum as detected by PCR in the trachea (P < 0.05) and conjunctiva (P < 0.01). Sinusitis was also associated with pasteurella cultured from the sinus (P < 0.05), antibody to avian pneumovirus (APV) (P < 0.01) and avian coronaviruses as detected by reverse-transcriptase PCR (P < 0.05); there was no association between disease and APV as detected by PCR. Avian coronaviruses were the most common infectious agents detected. They were genetically close to infectious bronchitis virus (IBV) but differed in their gene sequence from all the serotypes of IBV previously identified in domestic fowl, and serological tests with six known IBV types showed little cross reactivity. Mycoplasma species other than M gallisepticum were cultured in 18 batches of pheasants but, with the exception of Mycoplasma gallinaceum, were not associated with disease.
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PMID:Infectious agents associated with respiratory disease in pheasants. 1205 35

rpoB sequences encoding the beta-subunit of RNA polymerase were determined in 26 Mycoplasma species for phylogenetic study. The portion of rpoB DNA used in this study showed a high degree of variation in terms of size and sequence among species. The rpoB phylogenies inferred from amino acid and nucleotide sequences were used to divide the mycoplasmas into two groups, a 'pneumoniae group' and a 'hominis group', which was consistent with the result from 16S rDNA sequence analysis. However, phylogenetic relationships within these groups differed in the two gene trees, which were supported by the incongruence length difference (ILD) test. This indicates that multiple gene sequences must be applied to infer accurate phylogenetic relationships among the mycoplasmas. The rpoB sequence, and especially the deduced amino acid sequence, offers a good alternative marker.
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PMID:Use of rpoB sequences for phylogenetic study of Mycoplasma species. 1455 26

Gene sequences of small portions of the genome are often used for premature detailed taxonomic changes, neglecting polyphasic taxonomy, which should also consider phenotypical characteristics. Three examples are given: (i) Recently, members of the genera Eperythrozoon and Haemobartonella have been moved, correctly so, from the Rickettsiales to the Mycoplasmatales, but were assigned to the genus Mycoplasma, mostly on the basis of 16S rRNA sequence analysis. Not only is the 16S rRNA sequence similarity between 'classical' Mycoplasma and these species of Eperythrozoon and Haemobartonella less than that between some other well-recognised bacterial genera, but their biological differences amply justify their classification in different genera of the Mycoplasmatales. Furthermore, the move creates considerable confusion, as it necessitates new names for some species, with more confusion likely to come when the 16S rRNA sequences of the type species of Eperythrozoon, a name which has priority over Mycoplasma, will be analysed. (ii) In the Rickettsiales, members of the genera Anaplasma, Ehrlichia, Cowdria, Neorickettsia and Wolhbachia are so closely related phylogenetically on the basis of 16S rRNA sequences, and for some also of groESL operon sequences, that they have recently been fused, correctly so, into one family, the Anaplasmataceae, while the tribes Ehrlichieae and Wolbachieae have been abolished. Sequence diversity within the 'classical' genus Ehrlichia has led to classifying E. phagocytophila (including E. equi and the agent of human granulocytic ehrlichiosis), E. platys and E. bovis in the genus Anaplasma, while others have been retained in Ehrlichia, which also includes Cowdria ruminantium. E. sennetsu and E. risticii have been transferred to the genus Neorickettsia. 16S rRNA and GroEL sequences of 'classical' Anaplasma and some members of 'classical' Ehrlichia do show a close relationship, but differences in citrate synthase gene sequences, the GC content of this gene, and sequences of the gene encoding the beta-subunit of RNA polymerase, not to speak of the phenotypical differences, do not justify the fusion into one genus. Because of the phylogenetical diversity in Ehrlichia it is recommended that a new genus name be created for the E. phagocytophila genogroup (and E. platys and E. bovis). (iii) One of the conclusions of studies on the phylogeny of ticks of the subfamilies Rhipicephalinae and Hyalomminae, based on nucleotide sequences from 12S rRNA, cytochrome c oxidase I, the internal transcribed spacer 2, 18S rRNA, as well as morphological characters, is that Boophilus should be considered as a subgenus of Rhipicephalus. While Boophilus and Rhipicephalus are undoubtedly close, the obviously important morphological and biological differences between the genera Rhipicephalus and Boophilus are thus overruled by similarities in the sequences of a number of genes and this leads to considerable confusion. Polyphasic taxonomy amply justifies maintaining Boophilus as a separate genus, phylogenetically near to Rhipicephalus. This note is a plea for a cautious and balanced approach to taxonomy, taking into account molecular genotypical information, as far as is possible from different genes, as well as phenotypical characteristics.
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PMID:On molecular taxonomy: what is in a name? 1517 35


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