The gag precursor contains a specific HIV-1 protease cleavage site between the NC (P7) and P1 proteins.
The predicted protease cleavage site (p7/p1; [J. Virol. 66 (1992) 1856-1865]) within the nucleocapsid precursor protein (p15) of human immunodeficiency virus, type 1, was confirmed using an in vitro assay employing recombinant HIV-1 protease and a chemically synthesized 72 amino acid polypeptide containing the p7 and p1 protein domains of the native gag polyprotein. The cleavage occurred between amino acid 55 (N) and amino acid 56 (F) of the polypeptide, as determined by N-terminal sequencing. The hydrolysis was optimal at pH 6.0 and at high salt concentration. The kinetic parameters Km, kcat and kcat/Km were 99 microM (+/- 8), 0.152 s-1 (+/- 0.002) and 1.56 mM-1.s-1 (+/- 0.11), respectively. Reconstituted as well as denatured polypeptides were cleaved at approximately the same rate, demonstrating that the conformation of the p7 protein, as a result of the Zn(2+)-binding, had no significant effect on the rate of hydrolysis of the p7/p1 cleavage.
Wondrak
E M
EM
Unité de Recherches sur les Retrovirus et Maladies Associées, INSERM U 322, Marseille, France.
Louis
J M
JM
de Rocquigny
H
H
Chermann
J C
JC
Roques
B P
BP
D000595:Amino Acid Sequence; D001665:Binding Sites; D036022:Capsid Proteins; D002851:Chromatography, High Pressure Liquid; D015683:Gene Products, gag / Q000378:metabolism*; D016333:HIV Protease / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000378:metabolism*; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus
D036022:Capsid Proteins; D015683:Gene Products, gag; C071884:NCP7 protein, Human immunodeficiency virus 1; D011498:Protein Precursors; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus; C083868:gag protein p1, Human immunodeficiency virus; C059767:p55 gag precursor protein, Human immunodeficiency virus 1; D016333:HIV Protease