Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The genomic organization of Bordetella pertussis strains has been examined by using a new method. This method does not depend on the prior determination of a restriction map of the bacterial chromosome but is based on the ability to measure directly the distance between two genes. This is accomplished through the integration at each gene of a suicide vector containing a cleavage site for the intron-encoded endonuclease I-SceI, which is not otherwise found in the chromosome. Integration is mediated by homologous recombination between the chromosomal and cloned plasmid copies of a gene of interest. Digestion with I-SceI gives rise to a fragment the size of which represents the distance between the two genes. Multiple pairwise determinations within a set of genes provide sufficient information to derive a map of the relative gene positions. Mapping a set of 11 to 13 genes for five strains of B. pertussis and one strain of B. parapertussis revealed extensive divergence of gene order between B. pertussis Tohama I, B. pertussis 18-323, and B. parapertussis ATCC 15311. Less extensive divergence of gene order was observed between B. pertussis Tohama I and B. pertussis Tohama III, BP165, and Wellcome 28, with most of the observed differences explainable by large inversions.
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PMID:Genomic fluidity of Bordetella pertussis assessed by a new method for chromosomal mapping. 929 40

Background: The exosome complex plays key roles in RNA processing and degradation in Eukaryotes and Archaea. Outstanding structural studies identified multiple pathways for RNA substrates into the exosome in vitro, but identifying the pathway followed by individual RNA species in vivo remains challenging. Methods: We attempted to address this question using RNase protection. In vivo RNA-protein crosslinking (CRAC) was applied to the exosome component Rrp44/Dis3, which has both endonuclease and exonuclease activity. During CRAC, the exosome was purified under native conditions and subjected to RNase digestion, prior to protein denaturation and cDNA cloning. The resulting high-throughput sequence reads were stratified by length of the cDNA sequence. This should reflect RNA fragment lengths, and therefore the RNA region that was protected by exosome binding. We anticipated major read lengths of ~30nt and ~10nt, reflecting the "central channel" and "direct access" routes to the Rrp44 exonuclease active site observed in vitro. Results: Unexpectedly, no clear peak was observed at 30nt, whereas a broad peak was seen around 20nt. The expected ~10nt peak was seen, and showed strong elevation in strains lacking exonuclease activity. Unexpectedly, this peak was suppressed by point mutations in the Rrp44 endonuclease active site. This indicates that the short fragments are degraded by the exonuclease activity of Rrp44, but also suggests that at least some may be generated by endonuclease activity. Conclusions: The absence of 30nt protected fragments may reflect obligatory binding of cofactors at the entrance to the exosome central channel in vivo. The presence of ~20nt fragments apparently indicates an access route not yet reported from in vitro studies. Confident mapping of 10nt reads is challenging, but they are clearly derived from a subset of exosome targets. In particular, pre-rRNA species, which are major exosome targets, are strongly disfavored for the generation of short reads.
Wellcome Open Res 2017
PMID:RNA substrate length as an indicator of exosome interactions in vivo. 2874 21

The simplicity and the versatility of clustered regularly interspaced short palindromic repeats/CRISPR-associated protein (CRISPR-Cas) systems have enabled the genetic modification of virtually every organism and offer immense therapeutic potential for the treatment of human disease. Although these systems may function efficiently within eukaryotic cells, there remain concerns about the accuracy of Cas endonuclease effectors and their use for precise gene editing. Recently, two independent reports investigating the editing accuracy of the CRISPR-Cas9 system were published by separate groups at the Wellcome Sanger Institute; our study-Iyer and colleagues [1]-defined the landscape of off-target mutations, whereas the other by Kosicki and colleagues [2] detailed the existence of on-target, potentially deleterious deletions. Although both studies found evidence of large on-target CRISPR-induced deletions, they reached seemingly very different conclusions.
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PMID:Collateral damage and CRISPR genome editing. 3087 Apr 31