title: Structural plasticity and in vivo activity of Cas1 from the type I-F CRISPR-Cas system.
abstract: CRISPR-Cas systems are adaptive immune systems in prokaryotes that provide protection against viruses and other foreign DNA. In the adaptation stage, foreign DNA is integrated into CRISPR (clustered regularly interspaced short palindromic repeat) arrays as new spacers. These spacers are used in the interference stage to guide effector CRISPR associated (Cas) protein(s) to target complementary foreign invading DNA. Cas1 is the integrase enzyme that is central to the catalysis of spacer integration. There are many diverse types of CRISPR-Cas systems, including type I-F systems, which are typified by a unique Cas1-Cas2-3 adaptation complex. In the present study we characterize the Cas1 protein of the potato phytopathogen Pectobacterium atrosepticum, an important model organism for understanding spacer acquisition in type I-F CRISPR-Cas systems. We demonstrate by mutagenesis that Cas1 is essential for adaptation in vivo and requires a conserved aspartic acid residue. By X-ray crystallography, we show that although P. atrosepticum Cas1 adopts a fold conserved among other Cas1 proteins, it possesses remarkable asymmetry as a result of structural plasticity. In particular, we resolve for the first time a flexible, asymmetric loop that may be unique to type I-F Cas1 proteins, and we discuss the implications of these structural features for DNA binding and enzymatic activity.
authors.affiliation: Department of Microbiology and Immunology, University of Otago, PO Box 56, Dunedin 9054, New Zealand Department of Biochemistry, University of Otago, PO Box 56, Dunedin 9054, New Zealand.
authors.forename: Max E
authors.initials: ME
authors.lastname: Wilkinson
authors.affiliation: Department of Microbiology and Immunology, University of Otago, PO Box 56, Dunedin 9054, New Zealand Department of Biochemistry, University of Otago, PO Box 56, Dunedin 9054, New Zealand.
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Nakatani
authors.affiliation: Department of Microbiology and Immunology, University of Otago, PO Box 56, Dunedin 9054, New Zealand.
authors.forename: Raymond H J
authors.initials: RH
authors.lastname: Staals
authors.affiliation: Department of Microbiology and Immunology, University of Otago, PO Box 56, Dunedin 9054, New Zealand.
authors.forename: Sebastian N
authors.initials: SN
authors.lastname: Kieper
authors.affiliation: Department of Biochemistry, University of Otago, PO Box 56, Dunedin 9054, New Zealand.
authors.forename: Helen K
authors.initials: HK
authors.lastname: Opel-Reading
authors.affiliation: Department of Microbiology and Immunology, University of Otago, PO Box 56, Dunedin 9054, New Zealand.
authors.forename: Rebecca E
authors.initials: RE
authors.lastname: McKenzie
authors.affiliation: Department of Microbiology and Immunology, University of Otago, PO Box 56, Dunedin 9054, New Zealand peter.fineran@otago.ac.nz kurt.krause@otago.ac.nz.
authors.forename: Peter C
authors.initials: PC
authors.lastname: Fineran
authors.affiliation: Department of Biochemistry, University of Otago, PO Box 56, Dunedin 9054, New Zealand peter.fineran@otago.ac.nz kurt.krause@otago.ac.nz.
authors.forename: Kurt L
authors.initials: KL
authors.lastname: Krause
mesh_terms: D064130:CRISPR-Associated Proteins / Q000737:chemistry* / Q000378:metabolism*; D064113:CRISPR-Cas Systems / Q000502:physiology*; D003460:Crystallization; D018360:Crystallography, X-Ray; D004706:Endodeoxyribonucleases / Q000737:chemistry* / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D044043:Pectobacterium / Q000378:metabolism; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
chemical_list: D064130:CRISPR-Associated Proteins; D029968:Escherichia coli Proteins; D004706:Endodeoxyribonucleases; C557650:YgbT protein, E coli
keywords: Csy; adaptation; bacteriophages; horizontal gene transfer; plasmids; protein structure