The DNA-dependent and RNA-dependent DNA polymerase activities of the reverse transcriptases of human immunodeficiency viruses types 1 and 2.
We have constructed a series of plasmids which, when introduced into Escherichia coli, induce the overexpression of soluble wild-type and mutated forms of the reverse transcriptases (RTs) from human immunodeficiency viruses types 1 and 2 (HIV-1 and HIV-2, respectively). These proteins were analyzed previously for their RNA-dependent DNA polymerase (RDDP) and ribonuclease H (RNase H) activities. In the present study we assayed the different mutant RTs for their DNA-dependent DNA polymerase (DDDP) activity, employing an in situ polyacrylamide gel activity assay. The results indicate that both the RDDP and DDDP catalytic functions of HIV-1 RT mutants are affected similarly by mutations suggesting a high degree of overlap between the catalytic domains involved in both activities. Contrariwise, many of the HIV-2 RT mutants display no correlation between these two DNA polymerase activities, that is, the DDDP activity was not affected by the mutations introduced in the native enzyme in contrast to the RDDP activity. We were thus able to generate mutants of HIV-2 RT that unlike the wild-type RT, are capable of transcribing only DNA and not RNA. The disparity in mutational-catalytic relations between the two HIV-related RTs may reflect a possible difference in the structure and folding properties of the two proteins.
Shaharabany
M
M
Department of Cell Biology and Histology, Sackler School of Medicine, Tel Aviv University, Israel.
Hizi
A
A
D002384:Catalysis; D004252:DNA Mutational Analysis; D004279:DNA, Viral / Q000502:physiology*; D004259:DNA-Directed DNA Polymerase / Q000032:analysis* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D054303:HIV Reverse Transcriptase; D015497:HIV-1 / Q000201:enzymology* / Q000235:genetics; D015498:HIV-2 / Q000201:enzymology* / Q000235:genetics; D010957:Plasmids; D012367:RNA, Viral / Q000502:physiology*; D012194:RNA-Directed DNA Polymerase / Q000032:analysis* / Q000235:genetics
D004279:DNA, Viral; D012367:RNA, Viral; C072416:reverse transcriptase, Human immunodeficiency virus 2; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase; D004259:DNA-Directed DNA Polymerase