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  Modulating the Cascade architecture of a minimal Type I-F CRISPR-Cas system.

Gleditzsch, D., Müller-Esparza, H., Pausch, P., Sharma, K., Dwarakanath, S., Urlaub, H., et al. (2016). Modulating the Cascade architecture of a minimal Type I-F CRISPR-Cas system. Nucleic Acids Research, 44(12), 5872-5882. doi: 10.1093/nar/gkw469.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-002B-2E20-7 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-002C-AD76-D
Genre: Journal Article

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 Creators:
Gleditzsch, D., Author
Müller-Esparza, H., Author
Pausch, P., Author
Sharma, K.1, Author              
Dwarakanath, S., Author
Urlaub, H.1, Author              
Bange, G., Author
Randau, L., Author
Affiliations:
1Research Group of Bioanalytical Mass Spectrometry, MPI for Biophysical Chemistry, Max Planck Society, ou_578613              

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Free keywords: Diagnostic radiologic examination; roentgen rays; bacteriophage lambda; bacteriophages; gel chromatography; dna; immunity; mass spectrometry; infection; rna; complex; crispr
 Abstract: Shewanella putrefaciens CN-32 contains a single Type I-Fv CRISPR-Cas system which confers adaptive immunity against bacteriophage infection. Three Cas proteins (Cas6f, Cas7fv, Cas5fv) and mature CRISPR RNAs were shown to be required for the assembly of an interference complex termed Cascade. The Cas protein-CRISPR RNA interaction sites within this complex were identified via mass spectrometry. Additional Cas proteins, commonly described as large and small subunits, that are present in all other investigated Cascade structures, were not detected. We introduced this minimal Type I system in Escherichia coli and show that it provides heterologous protection against lambda phage. The absence of a large subunit suggests that the length of the crRNA might not be fixed and recombinant Cascade complexes with drastically shortened and elongated crRNAs were engineered. Size-exclusion chromatography and small-angle X-ray scattering analyses revealed that the number of Cas7fv backbone subunits is adjusted in these shortened and extended Cascade variants. Larger Cascade complexes can still confer immunity against lambda phage infection in E. coli. Minimized Type I CRISPR-Cas systems expand our understanding of the evolution of Cascade assembly and diversity. Their adjustable crRNA length opens the possibility for customizing target DNA specificity.

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Language(s): eng - English
 Dates: 2016-05-232016-07-08
 Publication Status: Published in print
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 Rev. Method: Peer
 Identifiers: DOI: 10.1093/nar/gkw469
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Title: Nucleic Acids Research
Source Genre: Journal
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Pages: - Volume / Issue: 44 (12) Sequence Number: - Start / End Page: 5872 - 5882 Identifier: -