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:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The chloroplast large subunit rRNA gene of Chlamydomonas eugametos and its 5' flanking region encoding tRNA(Ile) (GAU) and tRNA(Ala) (UGC) have been sequenced. The DNA sequence data along with the results of a detailed RNA analysis disclosed two unusual features of this green algal large subunit rRNA gene: (1) the presence of six group I introns (CeLSU.1-CeLSU.6) whose insertion positions have not been described previously, and (2) the presence of three short internal transcribed spacers that are post-transcriptionally excised to yield four rRNA species of 280, 52, 810 and 1720 nucleotides, positioned in this order (5' to 3') in the primary transcript. Together, these RNA species can assume a secondary structure that is almost identical to that proposed for the 23 S rRNA of Escherichia coli. All three internal transcribed spacers map to variable regions of primary sequence and/or potential secondary structure, whereas all six introns lie within highly conserved regions. The first three introns are inserted within the sequence encoding the 810 nucleotide rRNA species and map within domain II of the large subunit rRNA structure; the remaining introns, found in the sequence encoding the 1720 nucleotide rRNA species, lie within either domain IV or V, as is the case for all other large subunit rDNA introns that have been documented to date. CeLSU.5 and CeLSU.6 each contain a long open reading frame (ORF) of more than 200 codons. While the CeLSU.6 ORF is not related to any known ORFs, the CeLSU.5 ORF belongs to a family of ORFs that have been identified in Podospora and Neurospora mitochondrial group I introns. The finding that a polymorphic marker showing unidirectional gene conversion during crosses between C. eugametos and Chlamydomonas moewusii is located within the CeLSU.5 ORF makes it likely that this intron is a mobile element and that its ORF encodes a site-specific endonuclease promoting the transfer of the intron DNA sequence.
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PMID:Six group I introns and three internal transcribed spacers in the chloroplast large subunit ribosomal RNA gene of the green alga Chlamydomonas eugametos. 184 78

Splicing of tRNA precursors in extracts of Saccharomyces cerevisiae requires the action of two enzymes: a site specific endonuclease and a tRNA ligase. The tRNA ligase contains three distinct enzymatic activities: a polynucleotide kinase, a cyclic phosphodiesterase, and an RNA ligase. The polypeptide also has a high affinity pre-tRNA binding site based on its ability to form stable complexes with pre-tRNA substrates. To investigate the organization of functional enzymatic and binding elements within the polypeptide a series of defined tRNA ligase gene deletions were constructed and corresponding proteins were expressed in Escherichia coli as fusions with bacterial dihydrofolate reductase (DHFR). The DHFR/ligase derivative proteins were then efficiently purified by affinity chromatography. The complete ligase fusion protein retained enzymatic and binding activities which were unaffected by the presence of the DHFR segment. Examination of tRNA ligase deletion derivatives revealed that the amino-terminal region was required for adenylylation, while the carboxyl-terminal region was sufficient for cyclic phosphodiesterase activity. Deletions within the central region affected kinase activity. Pre-tRNA binding activity was not strictly correlated with a distinct enzymatic domain. A DHFR/ligase-derived protein lacking kinase activity efficiently joined tRNA halves. We postulate that this variant utilizes a novel RNA ligation mechanism.
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PMID:Deletion analysis of a multifunctional yeast tRNA ligase polypeptide. Identification of essential and dispensable functional domains. 185 Apr 8

Mitochondrial genes coding for some components of the protein synthetic apparatus in S. douglasii have been studies in detail. A region containing stretches of high homology to the S. cerevisiae tRNA synthesis locus (TSL) and the tRNA(fmet) gene has been identified and sequenced. The organization of this region was very similar to that present in S. cerevisiae, including the presence of a possible transcription starting signal. The S. douglasii TSL gene is shorter due to several deletions which, however, do not involve the regions coding for RNA domains know to be required for the catalytic activity of mitochondrial RNAse P. The S. douglasii LSU rRNA gene has been shown to contain a typical group I intron highly homologous to its S. cerevisiae counterpart, except for the absence of the open reading frame which in S. cerevisiae codes for I-SceI endonuclease.
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PMID:Mitochondrial genome of Saccharomyces douglasii: genes coding for components of the protein synthetic apparatus. 186 70

The biosynthesis of some mitochondrial enzymes requires contributions of both the mitochondrial and nuclear genomes. The ribonucleoprotein enzyme Ribonuclease P (RNase P) is composed of a mitochondrial encoded RNA and nuclear coded protein in many yeasts, including C. glabrata. We have determined that there are at least two sites of transcription initiation that contribute to the expression of the mitochondrial RNase P RNA. A nonanucleotide promoter sequence is located upstream of the initiator tRNA while the other site of initiation of transcription is at an undetermined upstream site. An analysis of the transcripts from the region of the RNase P gene demonstrates directly that the RNase P RNA is present in large primary transcripts and located between the precursors to the initiator tRNAf(Met) and tRNA(Pro) genes. Thus this enzyme subunit is synthesized with some of its substrate tRNAs. An activity with cleavage site specificity like a previously described endonuclease that cleaves near the 3' end of tRNAs, RNase P activity and one or more additional endonucleases or exonucleases not described previously are required to convert the primary transcript to its final functional RNAs.
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PMID:RNase P RNA in Candida glabrata mitochondria is transcribed with substrate tRNAs. 195 82

The mitochondrial DNA (mtDNA) sequence variation of the South American Ticuna, the Central American Maya, and the North American Pima was analyzed by restriction-endonuclease digestion and oligonucleotide hybridization. The analysis revealed that Amerindian populations have high frequencies of mtDNAs containing the rare Asian RFLP HincII morph 6, a rare HaeIII site gain, and a unique AluI site gain. In addition, the Asian-specific deletion between the cytochrome c oxidase subunit II (COII) and tRNA(Lys) genes was also prevalent in both the Pima and the Maya. These data suggest that Amerindian mtDNAs derived from at least four primary maternal lineages, that new tribal-specific variants accumulated as these mtDNAs became distributed throughout the Americas, and that some genetic variation may have been lost when the progenitors of the Ticuna separated from the North and Central American populations.
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PMID:Amerindian mitochondrial DNAs have rare Asian mutations at high frequencies, suggesting they derived from four primary maternal lineages. 196 8

Yeast tRNA(Trp)-encoding genes have been identified by Southern-blot analysis of chromosomal DNA. Seven copies of this gene are detected in blots of both restriction endonuclease digests and intact chromosomal DNA. Electrophoretic mapping of tDNA(Trp) indicates one copy is located on each of chromosomes X, XI, XIII, and XVI. The remaining three copies are localized to chromosomes VII and/or XV. Three different yeast strains gave identical results indicating this multi-gene family is relatively stable.
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PMID:Copy number and chromosomal location of Saccharomyces cerevisiae tRNA(Trp)-encoding genes. 202 21

Ribonuclease P is the endonuclease that removes the leader fragments from the 5'-ends of precursor tRNAs. The enzyme isolated from eubacteria contains a catalytic RNA subunit. RNAs also copurify with eukaryotic RNase P, although catalysis by those RNAs has not been demonstrated. This paper reports the isolation and characterization of ribonuclease P from the thermoacidophilic archaebacterium Sulfolobus solfataricus. Archaebacteria are a primary evolutionary lineage, distinct from both eukaryotes and eubacteria. Ribonuclease P of S. solfataricus has reaction component requirements and a Km for substrate tRNA (2.5 X 10(-7) M) that are roughly similar to those reported for eubacterial and eukaryotic ribonuclease P. The temperature optimum for the reaction is 77 degrees C, reflecting the thermophilic character of the organism. The enzyme activity is not affected by treatment with micrococcal nuclease, suggesting that there is no RNA subunit or that it is protected from nuclease action. The density of the enzyme in cesium sulfate equilibrium density gradients is 1.27 g/ml, which is similar to that of protein. However, several RNAs between 200 and 400 nucleotides in size copurify with the enzyme activity on the density gradients, and one of them remains after micrococcal nuclease treatment. These properties of the S. solfataricus enzyme are compared with those of ribonuclease P from eukaryotes and eubacteria.
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PMID:Characterization of ribonuclease P from the archaebacterium Sulfolobus solfataricus. 211 85

Inhibition of an RNA processing reaction after treatment with the Ca2(+)-dependent micrococcal nuclease (MN) is often used as a criterion for the presence of a required RNA or ribonucleoprotein component in the system. Following MN digestion, the nuclease is inactivated with EGTA and radiolabeled substrate is added to assay for remaining RNA processing activity. We found previously that inhibition of RNA processing by MN need not involve RNA hydrolysis: EGTA-inactivated MN can suppress RNA processing if the assay is performed in the absence of carrier RNA. We now demonstrate both by native gel electrophoresis and by nitrocellulose filter retention that EGTA-inactivated MN forms a complex with free RNA which can be dissociated by addition of synthetic polynucleotides or heparin. In the absence of Ca2+, nuclease binds to precursor tRNA with an apparent KD congruent to 1.4 x 10(-6) M, comparable to its reported affinity for DNA. In an assay for endonucleolytic tRNA maturation, inactivated MN bound to radiolabeled pre-tRNA physically blocks the sites of endonuclease cleavage and prevents tRNA processing. We call this phenomenon 'substrate masking'. Addition of excess carrier RNA competes with pre-tRNA for MN binding and restores normal processing.
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PMID:Substrate masking: binding of RNA by EGTA-inactivated micrococcal nuclease results in artifactual inhibition of RNA processing reactions. 212 40

The point mutation in the tRNA(Lys) gene of mitochondrial DNA (mtDNA) from patients with myoclonic epilepsy and ragged red fibers (MERRF) was quantitatively analyzed after digestion with the restriction endonuclease Nae I of the PCR amplified DNA. Since the point mutation is not part of a restriction site for a commonly available restriction endonuclease, the Nae I restriction site was introduced by PCR using a mispairing primer. The percentage of mutated mtDNA was determined in a few hairs of five members of an affected family by counting the radioactivity of the fragments after PCR amplification with labelled dATP.
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PMID:Identification of point mutations by mispairing PCR as exemplified in MERRF disease. 212 85

We identified and partially purified a phosphatase from crude extracts of Saccharomyces cerevisiae cells that can catalyze the last step of tRNA splicing in vitro. This phosphatase can remove the 2'-phosphate left over at the splice junction after endonuclease has removed the intron and ligase has joined together the two half-molecules. We suggest that this phosphatase is responsible for the completion of tRNA splicing in vivo, based primarily on its specificity for the 2'-phosphate of spliced tRNA and on the resistance of the splice junction 2'-phosphate to a nonspecific phosphatase. Removal of the splice junction 2'-phosphate from the residue adjacent to the anticodon is likely necessary for efficient expression of spliced tRNA. The phosphatase appears to be composed of at least two components which, together with endonuclease and ligase, can be used to reconstitute the entire tRNA-splicing reaction.
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PMID:A highly specific phosphatase from Saccharomyces cerevisiae implicated in tRNA splicing. 215 80


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