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Kex2p: a model for cellular endoprotease processing human immunodeficiency virus type 1 envelope glycoprotein
M Moulard1, T Achstetter, M P Kieny
1Laboratoire de Biochimie, CNRS URA 1455 - Ingénierie des Protéines, Faculté de Médecine Secteur Nord, Marseille, France.
European Journal of Biochemistry
|October 15, 1994
Summary
Yeast Kex2p protease can process the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein precursor (gp160). This finding suggests a functional similarity between yeast Kex2p and the cellular protease crucial for HIV-1 maturation.
Area of Science:
- Virology
- Molecular Biology
- Biochemistry
Background:
- Human immunodeficiency virus type 1 (HIV-1) requires endoproteolytic cleavage of its envelope glycoprotein precursor (gp160) for activation.
- Understanding the specific proteases involved is crucial for developing antiviral strategies.
Purpose of the Study:
- To investigate the in vitro processing of HIV-1 gp160 using yeast Kex2p as a model protease.
- To analyze the cleavage specificity of Kex2p on HIV-1 gp160 and a synthetic peptide mimic.
Main Methods:
- Coexpression of Kex2p and gp160 using recombinant vaccinia viruses.
- In vitro proteolysis assays using purified membranes from Saccharomyces cerevisiae overproducing Kex2p.
- Western blot analysis and N-terminal amino acid sequencing to identify cleavage products.
Main Results:
- Kex2p correctly processed HIV-1 gp160 into gp120 and gp41.
- Cleavage of recombinant gp160 occurred between Arg511 and Ala512, consistent with a dibasic cleavage site.
- Mutated gp160 and mature gp120 were not cleaved by Kex2p, indicating cleavage site specificity.
Conclusions:
- Yeast Kex2p demonstrates functional analogy to the cellular protease responsible for HIV-1 gp160 maturation.
- Kex2p effectively cleaves both synthetic peptides and recombinant gp160 at the conserved dibasic site.
- This study provides insights into the enzymatic mechanisms of HIV-1 glycoprotein processing.