Conformation and dynamics of the Gag polyprotein of the human immunodeficiency virus 1 studied by NMR spectroscopy

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 Delta Gag) by heteronuclear multidimensional NMR spectroscopy. In solution, Delta Gag exists in a dynamic equilibrium between monomeric and dimeric states. In the presence of nucleic acids and at low ionic strength Delta Gag assembles into immature virus-like particles. The structured domains of Delta Gag (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 Delta Gag 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 beta-hairpin upon cleavage at the matrix vertical bar capsid junction occurs five times faster than cleavage at the capsid vertical bar spacer peptide 1 vertical bar unction. Finally, nucleic acids interact with both nucleocapsid and matrix domains, and proteolytic processing at the spacer peptide 1 vertical bar nucleocapsid junction by HIV-1 protease is accelerated in the presence of single-stranded DNA.