title: Effects of temperature on the dynamic behaviour of the HIV-1 nucleocapsid NCp7 and its DNA complex.
abstract: The nucleocapsid protein NCp7 of human immunodeficiency virus type 1 (HIV-1) contains two highly conserved CCHC zinc fingers and is involved in many crucial steps of the virus life-cycle. A large number of physiological rôles of NCp7 involve its binding to single-stranded nucleic acid chains. Several solution structures of NCp7 and its complex with single-stranded RNA or DNA have been reported. We have investigated the changes in the dynamic behaviour experienced by the (12-53)NCp7 peptide upon DNA binding using (15)N heteronuclear relaxation measurements at 293 K and 308 K, and fluorescence spectroscopy. The relaxation data were interpreted using the reduced spectral density approach, which allowed the high-frequency motion, overall tumbling rates and the conformational exchange contributions to be characterized for various states of the peptide without using a specific motional model. Analysis of the temperature-dependent correlation times derived from both NMR and fluorescence data indicated a co-operative change of the molecular shape of apo (12-53)NCp7 around 303 K, leading to an increased hydrodynamic radius at higher temperatures. The binding of (12-53)NCp7 to a single-stranded d(ACGCC) pentanucleotide DNA led to a reduction of the conformational flexibility that characterized the apo peptide. Translational diffusion experiments as well as rotational correlation times indicated that the (12-53)NCp7/d(ACGCC) complex tumbles as a rigid object. The amplitudes of high-frequency motions were restrained in the complex and the occurrence of conformational exchange was displaced from the second zinc finger to the linker residue Ala30.
authors.affiliation: Laboratoire de Génomique Structurale, CNRS UPR 9004 ESBS, Bd Sébastien Brant, Illkirch, France.
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Ramboarina
authors.forename: Narayanan
authors.initials: N
authors.lastname: Srividya
authors.forename: R Andrew
authors.initials: RA
authors.lastname: Atkinson
authors.forename: Nelly
authors.initials: N
authors.lastname: Morellet
authors.forename: Bernard P
authors.initials: BP
authors.lastname: Roques
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Lefèvre
authors.forename: Yves
authors.initials: Y
authors.lastname: Mély
authors.forename: Bruno
authors.initials: B
authors.lastname: Kieffer
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002213:Capsid / Q000737:chemistry* / Q000378:metabolism*; D036022:Capsid Proteins; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D004058:Diffusion; D005454:Fluorescence Polarization; D015683:Gene Products, gag / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009038:Motion; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D018583:Pliability; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013696:Temperature; D013816:Thermodynamics; D014764:Viral Proteins; D016335:Zinc Fingers; D054301:gag Gene Products, Human Immunodeficiency Virus
chemical_list: D036022:Capsid Proteins; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D015683:Gene Products, gag; C071884:NCP7 protein, Human immunodeficiency virus 1; D010446:Peptide Fragments; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus