Quantitative in vitro assay for human immunodeficiency virus deoxyribonucleic acid integration.
An obligatory step of retroviral growth is the integration of a DNA copy of the viral RNA into the genomic DNA of the host. Recombinant human immunodeficiency virus type I (HIV-1) integrase (IN) expressed in Escherichia coli efficiently catalyzes the overall in vitro integration reaction, namely, the processing of the long terminal repeat (LTR) ends and the strand transfer reaction. Using the 3' end of synthetic oligonucleotides which match the termini of HIV-1 LTRs as substrate and supercoiled pSP65 DNA as the target, we describe an assay that is suitable for the enzymatic analysis of the integration and for testing candidate inhibitors of HIV IN protein.
Carteau
S
S
Laboratoire de Pharmacologie Moléculaire, CNRS URA 147, INSERM U 140, Institut Gustave Roussy, Villejuif, France.
Mouscadet
J F
JF
Goulaouic
H
H
Subra
F
F
Auclair
C
C
D001483:Base Sequence; D004254:DNA Nucleotidyltransferases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D019426:Integrases; D007700:Kinetics; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D016662:Virus Integration
D004279:DNA, Viral; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D004254:DNA Nucleotidyltransferases; D019426:Integrases