| Research Abstract |
A processing protease for the HIV-1 envelope glycoprotein gp160 precursor has been purified to homogeneity from the post-nuclear membrane fraction of human T4^+ lymphocyte clone. Most of the processing activity was found to be present in the fractions of endoplasmic reticulum and Golgi appratus of the cells. The purified enzyme has a monomeric structure with a molecular mass of 26*3kDa, as judged by gel-permeation liquid chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing and non-reducing conditions. The purified enzyme converted gp160 to gp120 and gp41, showing pH optimum of 6.5-7.0. Direct amino acid sequence of the amino terminus of the product gp41 revealed that the cleavage site of gp160 was between Arg^<511> and Ala^<512>. The enzyme activity was inhibited by trypsin-type protease inhibitors and by SH reagents, such as dithiothreitol, cysteine and N-acetyl L-cystein, but was not affected by CaCl2, MgCl2 or chelating agents. The properties of the purified enzyme are clearly distinct from those of processing proteases reported previously. Judging from its cleavage specificity and subcellular localization, this endopeptidase appears to be a processing enzyme for the HIV-1 gp160 precursor protein in human T cells. In cell culture system, low molecular weight protease inhibitors, such as diisopropylfluorophosphate, cystein and N-acetyl L-cystein, inhibited the processing of gp160 and the transportation of gp120 to the plasma membrane.
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