Use of chimeric human immunodeficiency virus types 1 and 2 reverse transcriptases for structure-function analysis and for mapping susceptibility to nonnucleoside inhibitors.
The human immunodeficiency virus type 1 and type 2 (HIV-1 and HIV-2) reverse transcriptases (RTs) are evolutionary related. To study the effect of homologous sequence replacements on polymerase function and to map the determinants of the lack of susceptibility of HIV-2 RT to nonnucleoside drugs, a series of chimeric HIV-1/HIV-2 RTs were constructed. Analysis of the chimeric RTs showed that wild-type levels of RNA-dependent DNA polymerase activity were retained when both finger and palm subdomains were exchanged as a unit between the two parental RTs. Analysis of enzymatically active chimeras for inhibition by the thiobenzimidazolone derivative TIBO R82150 showed that a segment of HIV-2 RT at 212-250, when placed in the HIV-1 RT context, conferred a 40-fold decrease in susceptibility to TIBO R82150. Site-directed mutagenesis of this segment found Tyr227 to be a key residue in this segment for the natural resistance of HIV-2 RT to TIBO R82150.
Yang
G
G
Department of Microbiology and Immunology, Albert Einstein College of Medicine, New York, New York, USA.
Song
Q
Q
Charles
M
M
Drosopoulos
W C
WC
Arnold
E
E
Prasad
V R
VR
D000596:Amino Acids / Q000502:physiology; D001483:Base Sequence; D001569:Benzodiazepines / Q000494:pharmacology*; D054303:HIV Reverse Transcriptase; D015497:HIV-1 / Q000201:enzymology*; D015498:HIV-2 / Q000201:enzymology*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D013329:Structure-Activity Relationship
D000596:Amino Acids; D007093:Imidazoles; D011993:Recombinant Fusion Proteins; D018894:Reverse Transcriptase Inhibitors; C062687:R 82150; D001569:Benzodiazepines; C072416:reverse transcriptase, Human immunodeficiency virus 2; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase