%0 Journal Article
%T Autoprocessing of human immunodeficiency virus type 1 protease miniprecursor fusions in mammalian cells
%A Liangqun Huang
%A Chaoping Chen
%J AIDS Research and Therapy
%D 2010
%I BioMed Central
%R 10.1186/1742-6405-7-27
%X We here describe a simple model system to directly examine HIV protease autoprocessing in transfected mammalian cells. A fusion precursor was engineered encoding GST fused to a well-characterized miniprecursor, consisting of the mature protease along with its upstream transframe region (TFR), and small peptide epitopes to facilitate detection of the precursor substrate and autoprocessing products. In HEK 293T cells, the resulting chimeric precursor undergoes effective autoprocessing, producing mature protease that is rapidly degraded likely via autoproteolysis. The known protease inhibitors Darunavir and Indinavir suppressed both precursor autoprocessing and autoproteolysis in a dose-dependent manner. Protease mutations that inhibit Gag processing as characterized using proviruses also reduced autoprocessing efficiency when they were introduced to the fusion precursor. Interestingly, autoprocessing of the fusion precursor requires neither the full proteolytic activity nor the majority of the N-terminal TFR region.We suggest that the fusion precursors provide a useful system to study protease autoprocessing in mammalian cells, and may be further developed for screening of new drugs targeting HIV protease autoprocessing.Human immunodeficiency virus 1 (HIV-1) is the causative pathogen of AIDS. The HIV protease is a virus-encoded enzyme absolutely required for virus propagation and infectivity. In the HIV infected cell, unspliced genomic RNA serves as mRNA for the synthesis of Gag and Gag-Pol polyproteins [1,2]. As part of the Gag-Pol polyprotein, the HIV protease is flanked at the N-terminus by a transframe region (TFR) and at the C-terminus by the reverse transcriptase [3,4]. The embedded protease has intrinsic but limited proteolytic activity [5,6] and the full activity is associated with the mature protease following its liberation from the precursor. Production of mature protease appears to be catalyzed by the Gag-Pol precursor itself serving as both the substrate
%U http://www.aidsrestherapy.com/content/7/1/27