Human immunodeficiency virus proteinase dimer as component of the viral polyprotein prevents particle assembly and viral infectivity.
Enzymatically active retroviral proteinases are dimers of identical polypeptide chains with a fold similar to that of other aspartic proteinases. Each polypeptide chain, encoded on one of the viral polyproteins, is less than half the size of cellular aspartic proteinases and contains only one of the two active-site aspartate residues. A plasmid was constructed to generate a genetically linked dimer of the proteinase (PR) of human immunodeficiency virus (HIV) type 1, composed of two copies of the PR sequence linked by a structurally flexible hinge region. The expression product was stable and active against HIV polyprotein substrates. Mutational analysis revealed that the linked dimer, and not multimers thereof, contained the proteolytic activity. Expression of the linked dimer as a component of a HIV polyprotein by in vitro translation gave rapid autocatalytic processing, whereas the wild-type polyprotein was stable on prolonged incubation. Transfection of HIV subviral or proviral constructs, containing the linked dimer of HIV PR, gave premature processing of the viral polyproteins, thus preventing particle formation and infectivity. Premature processing also led to increased cell toxicity.
Kräusslich
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HG
Institut für Virusforschung, Deutsches Krebsforschungszentrum, Heidelberg, Federal Republic of Germany.
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D015488:HIV Antigens / Q000378:metabolism; D016333:HIV Protease / Q000235:genetics / Q000648:ultrastructure*; D015497:HIV-1 / Q000201:enzymology* / Q000254:growth & development; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000378:metabolism*; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship; D014779:Virus Replication
D015488:HIV Antigens; D046911:Macromolecular Substances; D011506:Proteins; D011994:Recombinant Proteins; D016333:HIV Protease