Sequence requirements for encapsidation of deletion mutants and chimeras of human immunodeficiency virus type 1 Gag precursor into retrovirus-like particles.
Interacting domains in human immunodeficiency virus type 1 (HIV-1) Gag precursor (Pr55gag) expressed in recombinant baculovirus-infected cells were investigated by three different methods: (i) trans rescue and coencapsidation of C-terminal deletion (amber) Gag mutants and Gag chimeras into retrovirus-like particles in complementation experiments with HIV-1 wild-type (WT) Pr55gag, (ii) Gag-Gag interactions in vitro in Gag ligand affinity blotting assays, and (iii) quantitative immunoelectron microscopy of retrovirus-like Gag particles, using a panel of monoclonal antibodies to probe the epitope accessibility of encapsidated HIV-1 WT Pr55gag. Four discrete regions, within residues 210 to 241, 277 to 306 (major homology region), and 307 to 333 in the capsid (CA) protein and residues 358 to 374 at the CA-spacer peptide 2 (sp2) junction, were found to have a significant influence on Gag trans-packaging efficiency. A fifth region, within residues 375 to 426, overlapping the sp2-nucleocapsid (NC) protein junction and most of the NC, seemed to be essential for stable inter-Gag binding in vitro. The coincidence of the two regions from 358 to 374 and 375 to 426 with an immunologically silent domain in WT Gag particles suggested that they could participate in direct Gag interactions.
Carrière
C
C
Faculté de Médecine, Laboratoire de Virologie et Pathogénèse Moléculaires (CNRS URA-1487), Montpellier, France.
Gay
B
B
Chazal
N
N
Morin
N
N
Boulanger
P
P
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D003001:Cloning, Molecular; D015683:Gene Products, gag / Q000235:genetics*; D015497:HIV-1 / Q000235:genetics*; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D009154:Mutation; D011498:Protein Precursors / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics; D017384:Sequence Deletion; D018411:Spodoptera; D014771:Virion / Q000235:genetics* / Q000648:ultrastructure; D014779:Virus Replication
D015683:Gene Products, gag; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; C059767:p55 gag precursor protein, Human immunodeficiency virus 1