Active site mutagenesis of the AIDS virus protease and its alleviation by trans complementation.
Replacement of the putative active site Asp residue of cloned HIV-1 protease with Ala yields a molecule incapable of autocatalytic processing. Similarly, protease/reverse transcriptase and protease/reverse transcriptase/endonuclease polyproteins containing the same mutation accumulate as enzymatically inert polyproteins. Introduction of a second, wild-type, copy of protease in trans alleviates this defect, leading in the case of individually cloned protease to cleavage of the mutant protein, and with the polyprotein mutants to release of the reverse transcriptase and endonuclease polypeptides, the former of which recover enzymatic activity. In related experiments, a similar inhibition and trans-complementation of a genetically engineered gag--protease fusion protein was observed.
Le Grice
S F
SF
Central Research Units, F. Hoffmann-La Roche & Co. Ltd., Basel, Switzerland.
Mills
J
J
Mous
J
J
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004720:Endonucleases / Q000235:genetics; D015683:Gene Products, gag; D015497:HIV-1 / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D015151:Immunoblotting; D008969:Molecular Sequence Data; D009154:Mutation; D010447:Peptide Hydrolases / Q000235:genetics* / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012191:Retroviridae Proteins / Q000235:genetics
D015683:Gene Products, gag; D011993:Recombinant Fusion Proteins; D012191:Retroviridae Proteins; D012194:RNA-Directed DNA Polymerase; D004720:Endonucleases; D010447:Peptide Hydrolases