Base de dados : MEDLINE
Pesquisa : D12.776.212 [Categoria DeCS]
Referências encontradas : 426 [refinar]
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[PMID]:28456061
[Au] Autor:Richter F; Fonfara I; Gelfert R; Nack J; Charpentier E; Möglich A
[Ad] Endereço:Bayer AG, Pharmaceuticals, Protein Engineering and Assays, 50829 Köln, Germany. Electronic address: florian.richter@bayer.com.
[Ti] Título:Switchable Cas9.
[So] Source:Curr Opin Biotechnol;48:119-126, 2017 Dec.
[Is] ISSN:1879-0429
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Ever since its discovery, Cas9 from Streptococcus pyogenes has revolutionized biology by enabling analysis and engineering of genomes with unprecedented precision and ease. To fine-tune on-target effects and to mitigate adverse effects caused by untimely and off-target action of Cas9, strategies have been developed to control its activity at the post-translational stage via external trigger signals. Control is either achieved by modifying the Cas9 protein itself or its programmable RNA molecules. To date, switchable Cas9 variants responding to small ligands, light or temperature have been engineered. With these variants in hand, the regulation and modification of genomes can be accomplished in graded and ever more precise manner.
[Mh] Termos MeSH primário: Proteínas Associadas a CRISPR/metabolismo
Sistemas CRISPR-Cas
Regulação da Expressão Gênica
Engenharia Genética/métodos
Genoma Humano
[Mh] Termos MeSH secundário: Seres Humanos
Streptococcus pyogenes
[Pt] Tipo de publicação:JOURNAL ARTICLE; REVIEW
[Nm] Nome de substância:
0 (CRISPR-Associated Proteins)
[Em] Mês de entrada:1802
[Cu] Atualização por classe:180214
[Lr] Data última revisão:
180214
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170430
[St] Status:MEDLINE


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[PMID]:28985564
[Au] Autor:Guo TW; Bartesaghi A; Yang H; Falconieri V; Rao P; Merk A; Eng ET; Raczkowski AM; Fox T; Earl LA; Patel DJ; Subramaniam S
[Ad] Endereço:Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
[Ti] Título:Cryo-EM Structures Reveal Mechanism and Inhibition of DNA Targeting by a CRISPR-Cas Surveillance Complex.
[So] Source:Cell;171(2):414-426.e12, 2017 Oct 05.
[Is] ISSN:1097-4172
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Prokaryotic cells possess CRISPR-mediated adaptive immune systems that protect them from foreign genetic elements, such as invading viruses. A central element of this immune system is an RNA-guided surveillance complex capable of targeting non-self DNA or RNA for degradation in a sequence- and site-specific manner analogous to RNA interference. Although the complexes display considerable diversity in their composition and architecture, many basic mechanisms underlying target recognition and cleavage are highly conserved. Using cryoelectron microscopy (cryo-EM), we show that the binding of target double-stranded DNA (dsDNA) to a type I-F CRISPR system yersinia (Csy) surveillance complex leads to large quaternary and tertiary structural changes in the complex that are likely necessary in the pathway leading to target dsDNA degradation by a trans-acting helicase-nuclease. Comparison of the structure of the surveillance complex before and after dsDNA binding, or in complex with three virally encoded anti-CRISPR suppressors that inhibit dsDNA binding, reveals mechanistic details underlying target recognition and inhibition.
[Mh] Termos MeSH primário: Proteínas de Bactérias/química
Proteínas Associadas a CRISPR/química
Sistemas CRISPR-Cas
Microscopia Crioeletrônica
Pseudomonas aeruginosa/química
Pseudomonas aeruginosa/imunologia
[Mh] Termos MeSH secundário: Bacteriófagos/genética
Bacteriófagos/imunologia
Proteínas Associadas a CRISPR/imunologia
Proteínas Associadas a CRISPR/ultraestrutura
DNA Viral/química
Modelos Químicos
Modelos Moleculares
Complexos Multiproteicos/química
Pseudomonas aeruginosa/metabolismo
Pseudomonas aeruginosa/ultraestrutura
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (CRISPR-Associated Proteins); 0 (DNA, Viral); 0 (Multiprotein Complexes)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171115
[Lr] Data última revisão:
171115
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:171007
[St] Status:MEDLINE


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[PMID]:28911114
[Au] Autor:Liu T; Liu Z; Ye Q; Pan S; Wang X; Li Y; Peng W; Liang Y; She Q; Peng N
[Ad] Endereço:State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, 430070, P.R. China.
[Ti] Título:Coupling transcriptional activation of CRISPR-Cas system and DNA repair genes by Csa3a in Sulfolobus islandicus.
[So] Source:Nucleic Acids Res;45(15):8978-8992, 2017 Sep 06.
[Is] ISSN:1362-4962
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:CRISPR-Cas system provides the adaptive immunity against invading genetic elements in prokaryotes. Recently, we demonstrated that Csa3a regulator mediates spacer acquisition in Sulfolobus islandicus by activating the expression of Type I-A adaptation cas genes. However, links between the activation of spacer adaptation and CRISPR transcription/processing, and the requirement for DNA repair genes during spacer acquisition remained poorly understood. Here, we demonstrated that de novo spacer acquisition required Csa1, Cas1, Cas2 and Cas4 proteins of the Sulfolobus Type I-A system. Disruption of genes implicated in crRNA maturation or DNA interference led to a significant accumulation of acquired spacers, mainly derived from host genomic DNA. Transcriptome and proteome analyses showed that Csa3a activated expression of adaptation cas genes, CRISPR RNAs, and DNA repair genes, including herA helicase, nurA nuclease and DNA polymerase II genes. Importantly, Csa3a specifically bound the promoters of the above DNA repair genes, suggesting that they were directly activated by Csa3a for adaptation. The Csa3a regulator also specifically bound to the leader sequence to activate CRISPR transcription in vivo. Our data indicated that the Csa3a regulator couples transcriptional activation of the CRISPR-Cas system and DNA repair genes for spacer adaptation and efficient interference of invading genetic elements.
[Mh] Termos MeSH primário: Proteínas Arqueais/genética
Sistemas CRISPR-Cas
Reparo do DNA
DNA Arqueal/genética
Regulação da Expressão Gênica em Archaea
Sulfolobus/genética
Ativação Transcricional
[Mh] Termos MeSH secundário: Proteínas Arqueais/imunologia
Sequência de Bases
Proteínas Associadas a CRISPR/genética
Proteínas Associadas a CRISPR/imunologia
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas
DNA Helicases/genética
DNA Helicases/imunologia
DNA Polimerase II/genética
DNA Polimerase II/imunologia
DNA Arqueal/imunologia
Endodesoxirribonucleases/genética
Endodesoxirribonucleases/imunologia
Chaperonas Moleculares/genética
Chaperonas Moleculares/imunologia
Regiões Promotoras Genéticas
Alinhamento de Sequência
Homologia de Sequência do Ácido Nucleico
Sulfolobus/imunologia
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Archaeal Proteins); 0 (CRISPR-Associated Proteins); 0 (DNA, Archaeal); 0 (Molecular Chaperones); EC 2.7.7.- (DNA Polymerase II); EC 3.1.- (Endodeoxyribonucleases); EC 3.6.4.- (DNA Helicases)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171012
[Lr] Data última revisão:
171012
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170916
[St] Status:MEDLINE
[do] DOI:10.1093/nar/gkx612


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[PMID]:28893837
[Au] Autor:Burmistrz M; Rodriguez Martinez JI; Krochmal D; Staniec D; Pyrc K
[Ad] Endereço:Microbiology Department, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Cracow, Poland.
[Ti] Título:Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) RNAs in the Porphyromonas gingivalis CRISPR-Cas I-C System.
[So] Source:J Bacteriol;199(23), 2017 Dec 01.
[Is] ISSN:1098-5530
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:The CRISPR-Cas (clustered regularly interspaced short palindromic repeat-CRISPR-associated protein) system is unique to prokaryotes and provides the majority of bacteria and archaea with immunity against nucleic acids of foreign origin. CRISPR RNAs (crRNAs) are the key element of this system, since they are responsible for its selectivity and effectiveness. Typical crRNAs consist of a spacer sequence flanked with 5' and 3' handles originating from repeat sequences that are important for recognition of these small RNAs by the Cas machinery. In this investigation, we studied the type I-C CRISPR-Cas system in , a human pathogen associated with periodontitis, rheumatoid arthritis, cardiovascular disease, and aspiration pneumonia. We demonstrated the importance of the 5' handle for crRNA recognition by the effector complex and consequently activity, as well as secondary trimming of the 3' handle, which was not affected by modifications of the repeat sequence. , a clinically relevant Gram-negative, anaerobic bacterium, is one of the major etiologic agents of periodontitis and has been linked with the development of other clinical conditions, including rheumatoid arthritis, cardiovascular disease, and aspiration pneumonia. The presented results on the biogenesis and functions of crRNAs expand our understanding of CRISPR-Cas cellular defenses in and of horizontal gene transfer in bacteria.
[Mh] Termos MeSH primário: Sistemas CRISPR-Cas/genética
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética
Porphyromonas gingivalis/genética
RNA/genética
[Mh] Termos MeSH secundário: Proteínas Associadas a CRISPR/genética
Transferência Genética Horizontal/genética
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (CRISPR-Associated Proteins); 63231-63-0 (RNA)
[Em] Mês de entrada:1711
[Cu] Atualização por classe:171119
[Lr] Data última revisão:
171119
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170913
[St] Status:MEDLINE


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[PMID]:28890334
[Au] Autor:Billon P; Bryant EE; Joseph SA; Nambiar TS; Hayward SB; Rothstein R; Ciccia A
[Ad] Endereço:Department of Genetics and Development, Herbert Irving Comprehensive Cancer Center, Columbia University Medical Center, New York, NY 10032, USA.
[Ti] Título:CRISPR-Mediated Base Editing Enables Efficient Disruption of Eukaryotic Genes through Induction of STOP Codons.
[So] Source:Mol Cell;67(6):1068-1079.e4, 2017 Sep 21.
[Is] ISSN:1097-4164
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Standard CRISPR-mediated gene disruption strategies rely on Cas9-induced DNA double-strand breaks (DSBs). Here, we show that CRISPR-dependent base editing efficiently inactivates genes by precisely converting four codons (CAA, CAG, CGA, and TGG) into STOP codons without DSB formation. To facilitate gene inactivation by induction of STOP codons (iSTOP), we provide access to a database of over 3.4 million single guide RNAs (sgRNAs) for iSTOP (sgSTOPs) targeting 97%-99% of genes in eight eukaryotic species, and we describe a restriction fragment length polymorphism (RFLP) assay that allows the rapid detection of iSTOP-mediated editing in cell populations and clones. To simplify the selection of sgSTOPs, our resource includes annotations for off-target propensity, percentage of isoforms targeted, prediction of nonsense-mediated decay, and restriction enzymes for RFLP analysis. Additionally, our database includes sgSTOPs that could be employed to precisely model over 32,000 cancer-associated nonsense mutations. Altogether, this work provides a comprehensive resource for DSB-free gene disruption by iSTOP.
[Mh] Termos MeSH primário: Proteínas Associadas a CRISPR/genética
Sistemas CRISPR-Cas
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas
Códon de Terminação
Edição de Genes/métodos
Inativação Gênica
[Mh] Termos MeSH secundário: Animais
Arabidopsis/genética
Arabidopsis/metabolismo
Proteínas Associadas a CRISPR/metabolismo
Códon sem Sentido
Biologia Computacional
Enzimas de Restrição do DNA/genética
Enzimas de Restrição do DNA/metabolismo
Bases de Dados Genéticas
Regulação Fúngica da Expressão Gênica
Regulação Neoplásica da Expressão Gênica
Regulação da Expressão Gênica de Plantas
Células HEK293
Seres Humanos
Camundongos
Neoplasias/genética
Neoplasias/metabolismo
Polimorfismo de Fragmento de Restrição
RNA Guia/genética
RNA Guia/metabolismo
Ratos
Saccharomyces cerevisiae/genética
Saccharomyces cerevisiae/metabolismo
Transfecção
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (CRISPR-Associated Proteins); 0 (Codon, Nonsense); 0 (Codon, Terminator); 0 (RNA, Guide); EC 3.1.21.- (DNA Restriction Enzymes)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171116
[Lr] Data última revisão:
171116
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170912
[St] Status:MEDLINE


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[PMID]:28790199
[Au] Autor:El Refaey M; Xu L; Gao Y; Canan BD; Adesanya TMA; Warner SC; Akagi K; Symer DE; Mohler PJ; Ma J; Janssen PML; Han R
[Ad] Endereço:From the Department of Surgery, Davis Heart and Lung Research Institute, Biomedical Sciences Graduate Program, Biophysics Graduate Program (M.E.R., L.X., Y.G., T.M.A.A., J.M., R.H.) and Department of Physiology and Cell Biology, Davis Heart and Lung Research Institute (B.D.C., P.J.M., P.M.L.J.), The
[Ti] Título:In Vivo Genome Editing Restores Dystrophin Expression and Cardiac Function in Dystrophic Mice.
[So] Source:Circ Res;121(8):923-929, 2017 Sep 29.
[Is] ISSN:1524-4571
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:RATIONALE: Duchenne muscular dystrophy is a severe inherited form of muscular dystrophy caused by mutations in the reading frame of the dystrophin gene disrupting its protein expression. Dystrophic cardiomyopathy is a leading cause of death in Duchenne muscular dystrophy patients, and currently no effective treatment exists to halt its progression. Recent advancement in genome editing technologies offers a promising therapeutic approach in restoring dystrophin protein expression. However, the impact of this approach on Duchenne muscular dystrophy cardiac function has yet to be evaluated. Therefore, we assessed the therapeutic efficacy of CRISPR (clustered regularly interspaced short palindromic repeats)-mediated genome editing on dystrophin expression and cardiac function in mice after a single systemic delivery of recombinant adeno-associated virus. OBJECTIVE: To examine the efficiency and physiological impact of CRISPR-mediated genome editing on cardiac dystrophin expression and function in dystrophic mice. METHODS AND RESULTS: Here, we packaged SaCas9 (clustered regularly interspaced short palindromic repeat-associated 9 from ) and guide RNA constructs into an adeno-associated virus vector and systemically delivered them to neonates. We showed that CRIPSR-mediated genome editing efficiently excised the mutant exon 23 in dystrophic mice, and immunofluorescence data supported the restoration of dystrophin protein expression in dystrophic cardiac muscles to a level approaching 40%. Moreover, there was a noted restoration in the architecture of cardiac muscle fibers and a reduction in the extent of fibrosis in dystrophin-deficient hearts. The contractility of cardiac papillary muscles was also restored in CRISPR-edited cardiac muscles compared with untreated controls. Furthermore, our targeted deep sequencing results confirmed that our adeno-associated virus-CRISPR/Cas9 strategy was very efficient in deleting the ≈23 kb of intervening genomic sequences. CONCLUSIONS: This study provides evidence for using CRISPR-based genome editing as a potential therapeutic approach for restoring dystrophic cardiomyopathy structurally and functionally.
[Mh] Termos MeSH primário: Proteínas Associadas a CRISPR/genética
Sistemas CRISPR-Cas
Cardiomiopatias/terapia
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas
Distrofina/genética
Edição de Genes/métodos
Terapia Genética/métodos
Distrofia Muscular de Duchenne/terapia
Contração Miocárdica
Músculos Papilares/metabolismo
[Mh] Termos MeSH secundário: Animais
Proteínas Associadas a CRISPR/metabolismo
Cardiomiopatias/genética
Cardiomiopatias/metabolismo
Cardiomiopatias/fisiopatologia
Dependovirus/genética
Modelos Animais de Doenças
Distrofina/metabolismo
Éxons
Fibrose
Regulação da Expressão Gênica
Predisposição Genética para Doença
Vetores Genéticos
Sequenciamento de Nucleotídeos em Larga Escala
Camundongos Endogâmicos mdx
Distrofia Muscular de Duchenne/genética
Distrofia Muscular de Duchenne/metabolismo
Distrofia Muscular de Duchenne/fisiopatologia
Mutação
Músculos Papilares/patologia
Músculos Papilares/fisiopatologia
Fenótipo
RNA Guia/genética
RNA Guia/metabolismo
Recuperação de Função Fisiológica
Utrofina/genética
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (CRISPR-Associated Proteins); 0 (Dystrophin); 0 (RNA, Guide); 0 (Utrophin); 0 (apo-dystrophin 1)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171010
[Lr] Data última revisão:
171010
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170810
[St] Status:MEDLINE
[do] DOI:10.1161/CIRCRESAHA.117.310996


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[PMID]:28781236
[Au] Autor:Pausch P; Müller-Esparza H; Gleditzsch D; Altegoer F; Randau L; Bange G
[Ad] Endereço:LOEWE Center for Synthetic Microbiology (Synmikro) and Faculty of Chemistry, Philipps-University-Marburg, Hans-Meerwein-Strasse C07, 35043 Marburg, Germany.
[Ti] Título:Structural Variation of Type I-F CRISPR RNA Guided DNA Surveillance.
[So] Source:Mol Cell;67(4):622-632.e4, 2017 Aug 17.
[Is] ISSN:1097-4164
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:CRISPR-Cas systems are prokaryotic immune systems against invading nucleic acids. Type I CRISPR-Cas systems employ highly diverse, multi-subunit surveillance Cascade complexes that facilitate duplex formation between crRNA and complementary target DNA for R-loop formation, retention, and DNA degradation by the subsequently recruited nuclease Cas3. Typically, the large subunit recognizes bona fide targets through the PAM (protospacer adjacent motif), and the small subunit guides the non-target DNA strand. Here, we present the Apo- and target-DNA-bound structures of the I-Fv (type I-F variant) Cascade lacking the small and large subunits. Large and small subunits are functionally replaced by the 5' terminal crRNA cap Cas5fv and the backbone protein Cas7fv, respectively. Cas5fv facilitates PAM recognition from the DNA major groove site, in contrast to all other described type I systems. Comparison of the type I-Fv Cascade with an anti-CRISPR protein-bound I-F Cascade reveals that the type I-Fv structure differs substantially at known anti-CRISPR protein target sites and might therefore be resistant to viral Cascade interception.
[Mh] Termos MeSH primário: Proteínas de Bactérias/metabolismo
Proteínas Associadas a CRISPR/metabolismo
Sistemas CRISPR-Cas
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas
DNA/metabolismo
Endonucleases/metabolismo
Ácidos Nucleicos Heteroduplexes/metabolismo
RNA Guia/metabolismo
[Mh] Termos MeSH secundário: Proteínas de Bactérias/química
Proteínas de Bactérias/genética
Sítios de Ligação
Proteínas Associadas a CRISPR/química
Proteínas Associadas a CRISPR/genética
Cristalografia por Raios X
DNA/química
DNA/genética
Endonucleases/química
Endonucleases/genética
Escherichia coli/enzimologia
Escherichia coli/genética
Modelos Moleculares
Conformação de Ácido Nucleico
Ácidos Nucleicos Heteroduplexes/química
Ácidos Nucleicos Heteroduplexes/genética
Ligação Proteica
Conformação Proteica
Capuzes de RNA/metabolismo
RNA Guia/química
RNA Guia/genética
Shewanella putrefaciens/enzimologia
Shewanella putrefaciens/genética
Relação Estrutura-Atividade
[Pt] Tipo de publicação:COMPARATIVE STUDY; JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (CRISPR-Associated Proteins); 0 (Nucleic Acid Heteroduplexes); 0 (RNA Caps); 0 (RNA, Guide); 9007-49-2 (DNA); EC 3.1.- (Endonucleases)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170925
[Lr] Data última revisão:
170925
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170808
[St] Status:MEDLINE


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[PMID]:28781234
[Au] Autor:Yamano T; Zetsche B; Ishitani R; Zhang F; Nishimasu H; Nureki O
[Ad] Endereço:Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan.
[Ti] Título:Structural Basis for the Canonical and Non-canonical PAM Recognition by CRISPR-Cpf1.
[So] Source:Mol Cell;67(4):633-645.e3, 2017 Aug 17.
[Is] ISSN:1097-4164
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:The RNA-guided Cpf1 (also known as Cas12a) nuclease associates with a CRISPR RNA (crRNA) and cleaves the double-stranded DNA target complementary to the crRNA guide. The two Cpf1 orthologs from Acidaminococcus sp. (AsCpf1) and Lachnospiraceae bacterium (LbCpf1) have been harnessed for eukaryotic genome editing. Cpf1 requires a specific nucleotide sequence, called a protospacer adjacent motif (PAM), for target recognition. Besides the canonical TTTV PAM, Cpf1 recognizes suboptimal C-containing PAMs. Here, we report four crystal structures of LbCpf1 in complex with the crRNA and its target DNA containing either TTTA, TCTA, TCCA, or CCCA as the PAM. These structures revealed that, depending on the PAM sequences, LbCpf1 undergoes conformational changes to form altered interactions with the PAM-containing DNA duplexes, thereby achieving the relaxed PAM recognition. Collectively, the present structures advance our mechanistic understanding of the PAM-dependent, crRNA-guided DNA cleavage by the Cpf1 family nucleases.
[Mh] Termos MeSH primário: Proteínas de Bactérias/metabolismo
Proteínas Associadas a CRISPR/metabolismo
Sistemas CRISPR-Cas
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas
DNA/metabolismo
Endonucleases/metabolismo
Ácidos Nucleicos Heteroduplexes/metabolismo
RNA Guia/metabolismo
[Mh] Termos MeSH secundário: Acidaminococcus/enzimologia
Acidaminococcus/genética
Proteínas de Bactérias/química
Proteínas de Bactérias/genética
Sítios de Ligação
Proteínas Associadas a CRISPR/química
Proteínas Associadas a CRISPR/genética
Clostridiales/enzimologia
Clostridiales/genética
Cristalografia por Raios X
DNA/química
DNA/genética
Endonucleases/química
Endonucleases/genética
Escherichia coli/enzimologia
Escherichia coli/genética
Células HEK293
Seres Humanos
Modelos Moleculares
Conformação de Ácido Nucleico
Ácidos Nucleicos Heteroduplexes/química
Ácidos Nucleicos Heteroduplexes/genética
Ligação Proteica
Conformação Proteica
RNA Guia/química
RNA Guia/genética
Relação Estrutura-Atividade
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (CRISPR-Associated Proteins); 0 (Nucleic Acid Heteroduplexes); 0 (RNA, Guide); 9007-49-2 (DNA); EC 3.1.- (Endonucleases)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170925
[Lr] Data última revisão:
170925
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170808
[St] Status:MEDLINE


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[PMID]:28757251
[Au] Autor:Liu L; Li X; Ma J; Li Z; You L; Wang J; Wang M; Zhang X; Wang Y
[Ad] Endereço:Key Laboratory of RNA Biology, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
[Ti] Título:The Molecular Architecture for RNA-Guided RNA Cleavage by Cas13a.
[So] Source:Cell;170(4):714-726.e10, 2017 Aug 10.
[Is] ISSN:1097-4172
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Cas13a, a type VI-A CRISPR-Cas RNA-guided RNA ribonuclease, degrades invasive RNAs targeted by CRISPR RNA (crRNA) and has potential applications in RNA technology. To understand how Cas13a is activated to cleave RNA, we have determined the crystal structure of Leptotrichia buccalis (Lbu) Cas13a bound to crRNA and its target RNA, as well as the cryo-EM structure of the LbuCas13a-crRNA complex. The crRNA-target RNA duplex binds in a positively charged central channel of the nuclease (NUC) lobe, and Cas13a protein and crRNA undergo a significant conformational change upon target RNA binding. The guide-target RNA duplex formation triggers HEPN1 domain to move toward HEPN2 domain, activating the HEPN catalytic site of Cas13a protein, which subsequently cleaves both single-stranded target and collateral RNAs in a non-specific manner. These findings reveal how Cas13a of type VI CRISPR-Cas systems defend against RNA phages and set the stage for its development as a tool for RNA manipulation.
[Mh] Termos MeSH primário: Proteínas de Bactérias/química
Proteínas Associadas a CRISPR/química
Sistemas CRISPR-Cas
Leptotrichia/imunologia
[Mh] Termos MeSH secundário: Proteínas de Bactérias/ultraestrutura
Sequência de Bases
Proteínas Associadas a CRISPR/ultraestrutura
Leptotrichia/química
Leptotrichia/metabolismo
Leptotrichia/virologia
Modelos Moleculares
Processamento Pós-Transcricional do RNA
RNA Bacteriano/química
RNA Bacteriano/genética
RNA Bacteriano/ultraestrutura
RNA Guia/química
RNA Guia/genética
RNA Guia/ultraestrutura
RNA Viral/química
Difração de Raios X
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (CRISPR-Associated Proteins); 0 (RNA, Bacterial); 0 (RNA, Guide); 0 (RNA, Viral)
[Em] Mês de entrada:1708
[Cu] Atualização por classe:170816
[Lr] Data última revisão:
170816
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170801
[St] Status:MEDLINE


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[PMID]:28732207
[Au] Autor:Erard N; Knott SRV; Hannon GJ
[Ad] Endereço:Cancer Research UK Cambridge Institute, University of Cambridge, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK.
[Ti] Título:A CRISPR Resource for Individual, Combinatorial, or Multiplexed Gene Knockout.
[So] Source:Mol Cell;67(2):348-354.e4, 2017 Jul 20.
[Is] ISSN:1097-4164
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:We have combined a machine-learning approach with other strategies to optimize knockout efficiency with the CRISPR/Cas9 system. In addition, we have developed a multiplexed sgRNA expression strategy that promotes the functional ablation of single genes and allows for combinatorial targeting. These strategies have been combined to design and construct a genome-wide, sequence-verified, arrayed CRISPR library. This resource allows single-target or combinatorial genetic screens to be carried out at scale in a multiplexed or arrayed format. By conducting parallel loss-of-function screens, we compare our approach to existing sgRNA design and expression strategies.
[Mh] Termos MeSH primário: Proteínas Associadas a CRISPR/genética
Sistemas CRISPR-Cas
Endonucleases/genética
Inativação Gênica
Marcação de Genes/métodos
RNA Guia/genética
[Mh] Termos MeSH secundário: Algoritmos
Proteínas Associadas a CRISPR/metabolismo
Endonucleases/metabolismo
Biblioteca Gênica
Sequenciamento de Nucleotídeos em Larga Escala
Seres Humanos
Células K562
Aprendizado de Máquina
RNA Guia/metabolismo
Transfecção
[Pt] Tipo de publicação:COMPARATIVE STUDY; JOURNAL ARTICLE
[Nm] Nome de substância:
0 (CRISPR-Associated Proteins); 0 (RNA, Guide); EC 3.1.- (Endonucleases)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170926
[Lr] Data última revisão:
170926
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170722
[St] Status:MEDLINE



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