Autoprocessing: an essential step for the activation of HIV-1 protease.
Human immunodeficiency virus type 1 (HIV-1) expresses its structural and functional proteins within Gag-Pol precursor polyproteins. Specific proteolytic processing of the precursors by the viral protease is critical for the maturation and infectivity of viral particles. To observe the influence of autoprocessing on the activation of recombinant HIV-1 protease, we constructed different HIV-1 protease forms, with or without the Phe-Pro bond directly upstream of the protease domain, and expressed them in Escherichia coli systems. We found that the presence of a short upstream sequence of the protease domain, which could generate the original N-terminus of the protease by autoproteolysis of the Phe-Pro bond, resulted in processing of active protease, whereas for a wild-type protease extended only with the initiator methionine, the proteolytic activity was not recovered. Our results suggested that autoprocessing of the direct upstream sequence of the protease domain is an essential step for the activation of recombinant HIV-1 protease in the E. coli expression system. Expression of HIV-1 protease as fusion proteins revealed that the existence of a fusion portion increased the accumulation of expressed protease by affecting its homotypic dimer formation.
Wan
M
M
Department of Biochemistry, Faculty of Medicine, National University of Singapore.
Takagi
M
M
Loh
B N
BN
Xu
X Z
XZ
Imanaka
T
T
D001483:Base Sequence; D001665:Binding Sites; D015153:Blotting, Western; D017931:DNA Primers; D004789:Enzyme Activation; D004926:Escherichia coli / Q000235:genetics; D016333:HIV Protease / Q000378:metabolism*; D015497:HIV-1 / Q000201:enzymology*; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D017421:Sequence Analysis; D014764:Viral Proteins / Q000378:metabolism
D017931:DNA Primers; D010455:Peptides; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D016333:HIV Protease