title: Inter-domain orientation and motions in VAT-N explored by residual dipolar couplings and 15N backbone relaxation.
abstract: The N-terminal domain of VAT (Valosine-containing protein-like ATPase of Thermoplasma acidophilum), VAT-N (20.5 kDa), is considered to be the primary substrate-recognition site of the complex. The solution structure of VAT-N derived in our laboratory using conventionally obtained NMR restraints shows the existence of two equally sized sub-domains, VAT-Nn and VAT-Nc, together forming a kidney-shaped overall structure. The putative substrate-binding site of VAT-N involves free loops and a highly charged groove located on the surface of the protein. Alternatively, the opening of the cleft between the domains to accommodate substrate has been proposed to be part of the functional mechanism. We have used the residual dipolar couplings (RDCs) obtained in a bicelle medium to refine the structure of VAT-N. The long-range information available from RDCs both defines the sub-domain orientation and probes possible inter-domain motions. In addition, 15N backbone relaxation data were obtained and analysed within the model-free framework. Together, the data provides a refined structure with improved local geometry, but with the overall kidney shape intact. Further, the protein is rigid overall, with no evidence of inter-domain motions.
authors.affiliation: Department Chemie, Technische Universität München, Lichtenbergstrasse 4, 85747, Garching, Germany.
authors.forename: Mandar V
authors.initials: MV
authors.lastname: Deshmukh
authors.forename: Michael
authors.initials: M
authors.lastname: John
authors.forename: Murray
authors.initials: M
authors.lastname: Coles
authors.forename: Jürgen
authors.initials: J
authors.lastname: Peters
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Baumeister
authors.forename: Horst
authors.initials: H
authors.lastname: Kessler
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry*; D019843:Archaeal Proteins / Q000737:chemistry*; D001665:Binding Sites; D008958:Models, Molecular; D009587:Nitrogen Isotopes / Q000032:analysis; D019906:Nuclear Magnetic Resonance, Biomolecular; D017434:Protein Structure, Tertiary; D012996:Solutions; D013379:Substrate Specificity; D000074405:Valosin Containing Protein
chemical_list: D019843:Archaeal Proteins; D009587:Nitrogen Isotopes; D012996:Solutions; D000251:Adenosine Triphosphatases; D000074405:Valosin Containing Protein; C105134:vat protein, Thermoplasma acidophilum