Taylor, JE and Swiderska, A and Artero, JB and Callow, P and Kneale, G (2012) Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT). PLoS One, 7 (4). e35263 -?. ISSN 1932-6203

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Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT)..pdf - Published Version
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Abstract

Type I restriction-modification (RM) systems are comprised of two multi-subunit enzymes, the methyltransferase (∼160 kDa), responsible for methylation of DNA, and the restriction endonuclease (∼400 kDa), responsible for DNA cleavage. Both enzymes share a number of subunits. An engineered RM system, EcoR124I(NT), based on the N-terminal domain of the specificity subunit of EcoR124I was constructed that recognises the symmetrical sequence GAAN(7)TTC and is active as a methyltransferase. Here, we investigate the restriction endonuclease activity of R. EcoR124I(NT)in vitro and the subunit assembly of the multi-subunit enzyme. Finally, using small-angle neutron scattering and selective deuteration, we present a low-resolution structural model of the endonuclease and locate the motor subunits within the multi-subunit enzyme. We show that the covalent linkage between the two target recognition domains of the specificity subunit is not required for subunit assembly or enzyme activity, and discuss the implications for the evolution of Type I enzymes.

Item Type: Article
Uncontrolled Keywords: Base Sequence, Binding Sites, DNA, Deoxyribonucleases, Type I Site-Specific, Electrophoretic Mobility Shift Assay, Escherichia coli, Methyltransferases, Models, Molecular, Molecular Sequence Data, Neutron Diffraction, Protein Conformation, Protein Engineering, Protein Multimerization, Protein Structure, Tertiary, Protein Subunits, Scattering, Small Angle
Subjects: Q Science > QR Microbiology
Divisions: Faculty of Natural Sciences > School of Psychology
Related URLs:
Depositing User: Symplectic
Date Deposited: 15 Sep 2015 15:21
Last Modified: 30 Nov 2016 15:35
URI: http://eprints.keele.ac.uk/id/eprint/904

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