title: Conformation and dynamics of the Gag polyprotein of the human immunodeficiency virus 1 studied by NMR spectroscopy.
abstract: Assembly and maturation of the human immunodeficiency virus type 1 (HIV-1) are governed by the Gag polyprotein. Here we study the conformation and dynamics of a large HIV-1 Gag fragment comprising the matrix, capsid, spacer peptide 1 and nucleocapsid domains (referred to as ΔGag) by heteronuclear multidimensional NMR spectroscopy. In solution, ΔGag exists in a dynamic equilibrium between monomeric and dimeric states. In the presence of nucleic acids and at low ionic strength ΔGag assembles into immature virus-like particles. The structured domains of ΔGag (matrix, the N- and C-terminal domains of capsid, and the N- and C-terminal zinc knuckles of nucleocapsid) retain their fold and reorient semi-independently of one another; the linkers connecting the structural domains, including spacer peptide 1 that connects capsid to nucleocapsid, are intrinsically disordered. Structural changes in ΔGag upon proteolytic processing by HIV-1 protease, monitored by NMR in real-time, demonstrate that the conformational transition of the N-terminal 13 residues of capsid from an intrinsically disordered coil to a β-hairpin upon cleavage at the matrix|capsid junction occurs five times faster than cleavage at the capsid|spacer peptide 1 junction. Finally, nucleic acids interact with both nucleocapsid and matrix domains, and proteolytic processing at the spacer peptide 1|nucleocapsid junction by HIV-1 protease is accelerated in the presence of single-stranded DNA.
authors.affiliation: Laboratory of Chemical Physics and.
authors.forename: Lalit
authors.initials: L
authors.lastname: Deshmukh
authors.affiliation: Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520.
authors.forename: Rodolfo
authors.initials: R
authors.lastname: Ghirlando
authors.affiliation: Laboratory of Chemical Physics and mariusc@mail.nih.gov.
authors.forename: G Marius
authors.initials: GM
authors.lastname: Clore
mesh_terms: D000595:Amino Acid Sequence; D055592:Biophysical Phenomena; D002213:Capsid / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D016333:HIV Protease / Q000378:metabolism; D015497:HIV-1 / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D016624:Negative Staining; D017433:Protein Structure, Secondary; D059748:Proteolysis; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D054301:gag Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D011994:Recombinant Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus; D004247:DNA; D016333:HIV Protease; C514803:p16 protease, Human immunodeficiency virus 1
keywords: HIV-1 Gag; interdomain motion; proteolytic processing; real time NMR; residual dipolar couplings