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)

The molecular characterization of an additional DNA species (pAL2-1) which was identified previously in a long-lived extrachromosomal mutant (AL2) of Podospora anserina revealed that this element is a mitochondrial linear plasmid. pAL2-1 is absent from the corresponding wild-type strain, has a size of 8395 bp and contains perfect long terminal inverted repeats (TIRs) of 975 bp. Exonuclease digestion experiments indicated that proteins are covalently bound at the 5' termini of the plasmid. Two long, non-overlapping open reading frames, ORF1 (3,594 bp) and ORF2 (2847 bp), have been identified, which are located on opposite strands and potentially encode a DNA and an RNA polymerase, respectively. The ORF1-encoded polypeptide contains three conserved regions which may be responsible for a 3'-5' exonuclease activity and the typical consensus sequences for DNA polymerases of the D type. In addition, an amino-acid sequence motif (YSRLRT), recently shown to be conserved in terminal proteins from various bacteriophages, has been identified in the amino-terminal part of the putative protein. According to these properties, this first linear plasmid identified in P. anserina shares all characteristics with invertrons, a group of linear mobile genetic elements.
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PMID:The linear mitochondrial plasmid pAL2-1 of a long-lived Podospora anserina mutant is an invertron encoding a DNA and RNA polymerase. 147 81

The plus strand of the L-A double-stranded RNA virus of Saccharomyces cerevisiae has two large open reading frames, ORF1, which encodes the major coat protein, and ORF2, which encodes a single-stranded RNA-binding protein having a sequence diagnostic of viral RNA-dependent RNA polymerases. ORF2 is expressed only as a Gag-Pol-type fusion protein with ORF1. We have constructed a plasmid which expresses these proteins from the yeast PGK1 promoter. We show that this plasmid can support the replication of the killer toxin-encoding M1 satellite virus in the absence of an L-A double-stranded RNA helper virus itself. This requires ORF2 expression, providing a potential in vivo assay for the RNA polymerase and single-stranded RNA-binding activities of the fusion protein determined by ORF2. ORF1 expression, like a host ski- mutation, can suppress the usual requirement of M1 for the MAK11, MAK18, and MAK27 genes and allow a defective L-A (L-A-E) to support M1 replication. These results suggest that expression of ORF1 from the vector makes the cell a ski- phenocopy. Indeed, expression of ORF1 in a wild-type killer makes it a superkiller, suggesting that a target of the SKI antiviral system may be the major coat protein.
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PMID:Expression of yeast L-A double-stranded RNA virus proteins produces derepressed replication: a ski- phenocopy. 198 95

An 8-kilobase HindIII fragment carrying the histidase gene (hutH) and its regulatory region (hutP), from the Bacillus subtilis histidine utilization (hut) operon, was cloned in the temperate bacteriophage phi 105. Histidine utilization was restored in a hutH1 mutant by the specialized transducing phage (phi 105hutH11). The histidase gene in phi 105hutH11 was inducible and was shown to be under catabolite repression. The nucleotide sequence of 3,932 base pairs including the hutH and hutP loci revealed three open reading frames (ORFs). The molecular weights of ORF1 and ORF2 proteins were calculated to be 16,576 (151 amino acid residues) and 55,675 (508 amino acid residues), respectively. Reverse transcriptase mapping experiments showed that the putative promoter for the hut operon could be recognized by RNA polymerase sigma 43. The transcript starts at an adenosine residue 32 base pairs upstream from the initiation codon of ORF1. hutH+-transforming activity was found in ORF2, indicating that ORF2 encoded the histidase. A hutP1 mutation was determined as a substitution of an amino acid in ORF1. By using a specialized transducing phage containing the wild-type ORF1 gene, it was demonstrated that the presence of ORF1 protein in trans was absolutely required for the induction of the hut operon in a hutP1 mutant. These data strongly suggested that ORF1 encodes a positive regulator of the hut operon.
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PMID:Cloning and nucleotide sequences of histidase and regulatory genes in the Bacillus subtilis hut operon and positive regulation of the operon. 245 13

By a chromosome walking strategy the DNA region from Methanococcus vannielii flanking the genes for protein synthesis elongation factor (EF) 1 alpha and EF-2 was cloned and sequenced. A gene organization of 5' - beta' - open reading frame (ORF) 1 - ORF2 - S12 - S7 - EF-2 - EF-1 alpha - S10 - ORF3 - ORF4 - 3' was found where beta', S12, S7, S10, EF-2, and EF-1 alpha represent gene products with sequences similar to the beta' subunit of RNA polymerase, ribosomal proteins S12, S7, and S10, and EF-G and EF-Tu from Escherichia coli, respectively. ORF1-4 represent gene products with no known eubacterial counterparts. Northern blot analysis of transcripts and nuclease S1 mapping showed that transcription initiates between beta' and ORF1 and terminates at the 3' side of the S10 gene and that the genes from ORF1 to S10 are cotranscribed. Apart from the presence of two additional ORFs, ORF1 and ORF2, and of the gene for S10, this organization is identical to that of the eubacterial "streptomycin operon." ORF1 displays sequence similarity to rat liver ribosomal protein L30 and may represent one of the "additional" ribosomal proteins of Methanococcus. The sequenced part of the beta' gene and the EF-2 and EF-1 alpha gene products from Methanococcus are more similar to their eukaryotic than to their eubacterial counterparts. It appears, therefore, that the genetic organization of the translational components resembles the situation in eubacteria, whereas their primary structures are more eukaryotic in nature.
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PMID:Organization and nucleotide sequence of a transcriptional unit of Methanococcus vannielii comprising genes for protein synthesis elongation factors and ribosomal proteins. 247 40

The complete nucleotide sequences of the comA and comB loci of Bacillus subtilis were determined. The products of these genes are required for the development of competence in B. subtilis and for the expression of late-expressing competence genes. The major 5' termini of both the comA and comB transcripts were determined. The inferred promoters of both comA and comB contained sequences that were similar to those found at the -10 and -35 regions of promoters that are used by sigma A-RNA polymerase, the primary form of this enzyme in vegetative cells. The comB gene was located approximately 3 kilobase pairs upstream of the comA gene and encoded a 409-amino-acid protein with a predicted molecular weight of 46,693. The comA locus contained two open reading frames (ORFs) and comB contained one ORF. The predicted amino acid sequence of the comA ORF1 gene product consisted of 214 amino acids, with an aggregate molecular weight of 24,132. The ORF1 product was required for competence, while ORF2, which was cotranscribed with ORF1 and encoded a predicted protein of 126 amino acids, was not. The predicted protein sequence of the comA ORF1 gene product was found to be similar to that of several members of the effector class of procaryotic signal transducers. The C-terminal portion of the predicted comA sequence contained a possible helix-turn-helix motif, which is characteristic of DNA-binding proteins. comA ORF1 was cloned on a multicopy plasmid and was shown to complement the competence-deficient phenotype caused by the comA124 insertion of Tn917lac. Also, the presence of comA ORF1 in multiple copies interfered with sporulation. Anti-peptide antibodies raised to the predicted product of comA ORF1 reacted strongly with a single protein band of about 24,000 daltons in immunoblots. The possible roles of multiple signal transduction systems in triggering the development of competence are discussed.
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PMID:Sequence and transcription mapping of Bacillus subtilis competence genes comB and comA, one of which is related to a family of bacterial regulatory determinants. 250 23

A series of Tn917lac insertions define the comG region of the Bacillus subtilis chromosome. comG mutants are deficient in competence and specifically in the binding of exogenous DNA. The genes included in the comG region are first expressed during the transition from the exponential to the stationary growth phase. From nucleotide sequence information, it was concluded that the comG locus contains seven open reading frames (ORFs), several of which overlap at their termini. High-resolution S1 nuclease mapping and primer extension were used to identify the 5' terminus of the comG mRNA. The sequence upstream from the comG start site closely resembled the consensus recognition sequence for the major B. subtilis vegetative RNA polymerase holoenzyme. Complementation analysis confirmed that the comG ORF1 protein is required for the ability of competent cultures to resolve into two populations with different cell densities on Renografin (E. R. Squibb & Sons, Princeton, N.J.) gradients, as well as for full expression of comE, another late competence locus. The predicted comG ORF1 protein showed significant similarity to the virB ORF11 protein from Agrobacterium tumefaciens, which is probably involved in T-DNA transfer. The N-terminal sequences of comG ORF3 and, to a lesser extent, the comG ORF4 and ORF5 proteins were similar to a class of pilin proteins from members of the genera Bacteroides, Pseudomonas, Neisseria, and Moraxella. All of the comG proteins except comG ORF1 possessed hydrophobic domains that were potentially capable of spanning the bacterial membrane. It is likely that these proteins are membrane associated, and they may comprise part of the DNA transport machinery. When present in multiple copies, a DNA fragment carrying the comG promoter was capable of inhibiting the development of competence as well as the expression of several late com genes, suggesting a role for a transcriptional activator in the expression of those genes.
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PMID:Nucleotide sequence and genetic organization of the Bacillus subtilis comG operon. 250 24

The complete nucleotide sequence has been determined of a 3635-bp region, extending from the HpaI site in traT, at F coordinate 90.3 kb, to beyond the end of traD, of the F sex factor plasmid of Escherichia coli K-12. This region contains the C-terminal coding part of traT and the entire traD gene. An open reading frame (ORF) of 2148 bp within the sequence confirms that traD encodes an 81.4-kDa cytoplasmic membrane protein. The TraD protein has several regions with an unusually high pI (greater than 10), suggesting that they may correspond to the DNA-binding domains. Several other ORFs were detected within the region including the gene (ORF1) for a 26.3-kDa protein and ORF2, probably corresponding to traI, which continues to the end of the sequence. An ORF for an 8.5-kDa protein preceded by an excellent promoter and ribosome-binding site is present in the region following traD but on the opposite strand. This promoter is thought to correspond to the major RNA polymerase binding site in this region, implying that traI does not have its own promoter. The lack of a typical terminator following traD and ORF1 and the translational coupling provided by overlapping stop and start codons is consistent with this conclusion.
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PMID:Nucleotide sequence of the traD region in the Escherichia coli F sex factor. 268 Jul 68

Directly upstream of the Lactococcus lactis subsp. cremoris Wg2 proteinase gene is an oppositely directed open reading frame (ORF1). The complete nucleotide sequence of ORF1, encoding a 33-kilodalton protein, was determined. A protein of approximately 32 kilodaltons was synthesized when ORF1 was expressed in Escherichia coli by using a T7 RNA polymerase-specific promoter. L. lactis subsp. lactis MG1363 transformants carrying the proteinase gene but lacking ORF1 were phenotypically proteinase deficient, unlike transformants carrying both the proteinase gene and ORF1. Synthesis and secretion of proteinase antigen by L. lactis could be detected with proteinase-directed monoclonal antibodies regardless of whether ORF1 was present. The requirement of ORF1 for proteinase activation was reflected in a reduction in the molecular weight of the secreted proteinase. Furthermore, deletion of the 130 C-terminal amino acids of the Wg2 proteinase prevented attachment of the enzyme to lactococcal cells.
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PMID:Identification of a gene required for maturation of an extracellular lactococcal serine proteinase. 270 18

Rabbit hemorrhagic disease virus, a positive-stranded RNA virus of the family Caliciviridae, encodes a trypsin-like cysteine protease as part of a large polyprotein. Upon expression in Escherichia coli, the protease releases itself from larger precursors by proteolytic cleavages at its N and C termini. Both cleavage sites were determined by N-terminal sequence analysis of the cleavage products. Cleavage at the N terminus of the protease occurred with high efficiency at an EG dipeptide at positions 1108 and 1109. Cleavage at the C terminus of the protease occurred with low efficiency at an ET dipeptide at positions 1251 and 1252. To study the cleavage specificity of the protease, amino acid substitutions were introduced at the P2, P1, and P1' positions at the cleavage site at the N-terminal boundary of the protease. This analysis showed that the amino acid at the P1 position is the most important determinant for substrate recognition. Only glutamic acid, glutamine, and aspartic acid were tolerated at this position. At the P1' position, glycine, serine, and alanine were the preferred substrates of the protease, but a number of amino acids with larger side chains were also tolerated. Substitutions at the P2 position had only little effect on the cleavage efficiency. Cell-free expression of the C-terminal half of the ORF1 polyprotein showed that the protease catalyzes cleavage at the junction of the RNA polymerase and the capsid protein. An EG dipeptide at positions 1767 and 1768 was identified as the putative cleavage site. Our data show that rabbit hemorrhagic disease virus encodes a trypsin-like cysteine protease that is similar to 3C proteases with regard to function and specificity but is more similar to 2A proteases with regard to size.
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PMID:3C-like protease of rabbit hemorrhagic disease virus: identification of cleavage sites in the ORF1 polyprotein and analysis of cleavage specificity. 747 37

Comparisons of the RNA polymerase and capsid sequences of small round structured viruses (SRSVs) have recently shown these are genetically diverse viruses which fall into two distinct groups. The genomes of two group I viruses, Southampton and Norwalk viruses have been characterized; however, similar data for the genetic group II SRSVs have not been available until now. We report here the complete genome sequence of a recent group II SRSV, Lordsdale virus. The Lordsdale virus genome is 7555 nt in length and has a similar organization to the group I SRSVs. The large ORF in the 5' half of the genome (5100 nt) is shorter than the group I SRSV ORF1 (5367 nt), but has the characteristic 2C helicase, 3C protease and 3D RNA polymerase enzyme motifs. ORF2, encoding the structural protein is of a similar size to the group I viruses but the small 3'-terminal ORF is significantly larger in group II. A highly conserved sequence of 28 nt was identified at the start of Lordsdale virus ORF1 and repeated at the start of ORF2. These conserved motifs are typical of the animal caliciviruses. Comparison of the 150 N-terminal amino acids in the ORF1 protein revealed little identity between the two SRSV genetic groups, reflecting the shorter ORF1 in the group II virus. Recombinant baculoviruses containing ORF2 and ORF3 sequences were constructed and used to express large quantities of the group II Lordsdale virus structural protein. The capsid protein formed virus-like particles by self assembly which resembled 'empty' SRSVs.
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PMID:Human enteric Caliciviridae: the complete genome sequence and expression of virus-like particles from a genetic group II small round structured virus. 756 76


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