title: Nucleotide-type chemical shift assignment of the encapsulated 40 kbp dsDNA in intact bacteriophage T7 by MAS solid-state NMR.
abstract: The icosahedral bacteriophage T7 is a 50 MDa double-stranded DNA (dsDNA) virus that infects Escherichia coli. Although there is substantial information on the physical and morphological properties of T7, structural information, based mostly on Raman spectroscopy and cryo-electron microscopy, is limited. Here, we apply the magic-angle spinning (MAS) solid-state NMR (SSNMR) technique to study a uniformly (13)C and (15)N labeled wild-type T7 phage. We describe the details of the large-scale preparation and purification of an isotopically enriched phage sample under fully hydrated conditions, and show a complete (13)C and a near-complete (15)N nucleotide-type specific assignment of the sugar and base moieties in the 40 kbp dsDNA of T7 using two-dimensional (13)C-(13)C and (15)N-(13)C correlation experiments. The chemical shifts are interpreted as reporters of a B-form conformation of the encapsulated dsDNA. While MAS SSNMR was found to be extremely useful in determining the structures of proteins in native-like environments, its application to nucleic acids has lagged behind, leaving a missing (13)C and (15)N chemical shift database. This work therefore expands the (13)C and (15)N database of real B-form DNA systems, and opens routes to characterize more complex nucleic acid systems by SSNMR.
authors.affiliation: School of Chemistry, Raymond and Beverly Sackler Faculty of Exact Sciences, Tel Aviv University, 69978, Ramat Aviv, Tel Aviv, Israel.
authors.forename: Gili
authors.initials: G
authors.lastname: Abramov
authors.forename: Amir
authors.initials: A
authors.lastname: Goldbourt
mesh_terms: D017123:Bacteriophage T7 / Q000737:chemistry*; D004279:DNA, Viral / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular; D009690:Nucleic Acid Conformation
chemical_list: D004279:DNA, Viral