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A high throughput assay for inhibitors of HIV‐1 protease Screening of microbial metabolites
Author(s) -
Sarubbi Edoardo,
Nolli M.Luisa,
Andronico Franca,
Stella Sergio,
Saddler Gerard,
Selva Enrico,
Siccardi Antonio,
Denaro Maurizio
Publication year - 1991
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(91)80164-x
Subject(s) - protease , hiv 1 protease , monoclonal antibody , fusion protein , recombinant dna , chemistry , cleavage (geology) , human immunodeficiency virus (hiv) , high throughput screening , biochemistry , protease inhibitor (pharmacology) , enzyme , tandem affinity purification , microbiology and biotechnology , antibody , chromatography , biology , virology , affinity chromatography , gene , antiretroviral therapy , paleontology , viral load , immunology , fracture (geology)
A novel method for discovery of HIV‐1 protease inhibitors in complex biological samples has been developed. The assay is based on two specific reagents: a recombinant protein constituted by a portion of the HIV‐1 Gag polyprotein comprising the p17–p24 cleavage site, fused to E. coli β‐galactoxidase, and a monoclonal antibody which binds the fusion protein in the Gag region. Binding occurs only if the fusion protein has not been cleaved by the HIV‐1 protease. The assay has been adapted for the screening of large numbers of samples in standard 96‐well microliter plates. Using this method about 12000 microbial fermentation broths have been tested and several HIV‐1 protease inhibitory activities have been detected. One of these has been studied in detail.