A high throughput assay for inhibitors of HIV-1 protease. Screening of microbial metabolites.
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 beta-galactosidase, 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 microtiter 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.
Sarubbi
E
E
Lepetit Research Center, MMDRI, Gerenzano VA, Italy.
Nolli
M L
ML
Andronico
F
F
Stella
S
S
Saddler
G
G
Selva
E
E
Siccardi
A
A
Denaro
M
M
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D015683:Gene Products, gag / Q000378:metabolism; D017320:HIV Protease Inhibitors; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D000911:Antibodies, Monoclonal; D015683:Gene Products, gag; D017320:HIV Protease Inhibitors; D011480:Protease Inhibitors; D011993:Recombinant Fusion Proteins