In vivo processing of Pr160gag-pol from human immunodeficiency virus type 1 (HIV) in acutely infected, cultured human T-lymphocytes.
The processing of the HIV-1 Pr160gag-pol precursor polyprotein was analyzed in freshly HIV-1-infected MT-4 cultured cells. Single intermediates of the processing cascade were characterized by immunoblotting using distinct antisera. A potent inhibitor of the HIV protease (PR), Ro 31-8959, was employed to block cleavage by the mature PR, thus allowing insights into initial stages of the gag-pol (auto)-catalytical processing. While most known gag-pol cleavages were blocked in the presence of the inhibitor, the cleavage site between the gag-NC and the pol-p6 domains was still cleaved even in presence of high amounts (1 microM) of inhibitor, leading to the accumulation of a novel 114-kDa polyprotein comprising p6-PR-RT-IN. In the absence of inhibitor no accumulation of p114 was observed. In inhibitor-treated, HIV-1-infected cells a p6-PR intermediate was also detected, indicating subsequent cleavage of the PR/RT scissile bond. These results demonstrate initial cleavage(s) of the gag-pol precursor hydrolyzed by a proteolytic activity different from the mature PR and indicate that p114 (p6-PR-RT-IN) and p6-PR intermediates could play an essential role in the PR activation process.
Lindhofer
H
H
Max von Pettenkofer-Institute, University of Munich, Germany.
von der Helm
K
K
Nitschko
H
H
D001665:Binding Sites; D002460:Cell Line; D004792:Enzyme Precursors / Q000378:metabolism; D015683:Gene Products, gag / Q000378:metabolism*; D016333:HIV Protease / Q000378:metabolism; D017320:HIV Protease Inhibitors / Q000494:pharmacology; D006801:Humans; D007546:Isoquinolines / Q000494:pharmacology; D011498:Protein Precursors / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011804:Quinolines / Q000494:pharmacology; D019258:Saquinavir; D013379:Substrate Specificity; D013601:T-Lymphocytes / Q000821:virology*; D054301:gag Gene Products, Human Immunodeficiency Virus; D054302:pol Gene Products, Human Immunodeficiency Virus
D004792:Enzyme Precursors; D015683:Gene Products, gag; D017320:HIV Protease Inhibitors; D007546:Isoquinolines; D011498:Protein Precursors; D011804:Quinolines; D054301:gag Gene Products, Human Immunodeficiency Virus; D054302:pol Gene Products, Human Immunodeficiency Virus; C091950:pr160 gag-pol precursor protein, Human immunodeficiency virus 1; D016333:HIV Protease; D019258:Saquinavir