Assembly and extracellular release of chimeric HIV-1 Pr55gag retrovirus-like particles.
The HIV-1 Pr55gag precursors were previously shown to assemble and bud from a variety of different cell types as noninfectious virus-like particles (VLPs) resembling immature HIV virions. The use of these VLPs as an immunogenic and autologous carrier component may allow the presentation of defined epitopes deduced from reading frames other than gag to the immune system, thereby avoiding the induction of adverse immune responses. In order to identify domains within Pr55gag that can be replaced by immunologically relevant epitopes without affecting its capacity to assemble into VLPs, we deleted three domains of a predicted high surface probability. Deletion of amino acids 211-241 within p24CA and amino acids 436-471 within the p6LI portion of Pr55gag had no effect on the assembly, ultrastructure, biophysical properties, and yields of mutant VLPs when expressed via recombinant vaccinia viruses in mammalian cells. Deletion of amino acids 99-154 overlapping the p17MA/p24CA cleavage site completely abolished the capacity of the gag polyprotein to form VLPs and led to a reduction of immature Pr55 VLPs released into the cell-culture supernatants when coexpressed with wild-type Pr55gag. In contrast, assembly and budding of chimeric VLPs could be demonstrated after replacing amino acids 211-241 and 436-471 by immunologically relevant epitopes derived from reading frames other than Pr55gag (e.g., V3 loop; CD4-binding-domain; nef-CTL epitope) or after fusion of these sequences to the carboxy terminus of Pr55gag. The importance of these data for the development of novel HIV candidate vaccines is discussed.
Wagner
R
R
Institut für med. Mikrobiologie, LMU-München, Federal Republic of Germany.
Deml
L
L
Fliessbach
H
H
Wanner
G
G
Wolf
H
H
D000595:Amino Acid Sequence; D001483:Base Sequence; D004252:DNA Mutational Analysis; D000939:Epitopes / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015683:Gene Products, gag / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015699:HIV Envelope Protein gp120 / Q000235:genetics / Q000276:immunology; D015497:HIV-1 / Q000235:genetics / Q000254:growth & development* / Q000276:immunology / Q000648:ultrastructure; D008969:Molecular Sequence Data; D009154:Mutation; D011498:Protein Precursors / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014616:Vaccinia virus / Q000235:genetics / Q000648:ultrastructure; D014771:Virion / Q000235:genetics / Q000254:growth & development* / Q000276:immunology / Q000648:ultrastructure
D000939:Epitopes; D015683:Gene Products, gag; D015699:HIV Envelope Protein gp120; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; C059767:p55 gag precursor protein, Human immunodeficiency virus 1