Mutational analysis of the ribonuclease H activity of human immunodeficiency virus 1 reverse transcriptase

Amnon Hizi*, Stephen H. Hughes, Miriam Shaharabany

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

77 Scopus citations


We have constructed a series of plasmids that, when introduced into Escherichia coli, induce the expression of high levels of either wild-type or mutated forms of the reverse transcriptase (RT) of human immunodeficiency virus type 1 (HIV-1). Mutant forms of RT that had been previously analyzed for their RNA-dependent DNA polymerse activity were tested for RNase H activity using an in situ polyacrylamide gel assay. Mutations affecting the RNase H are not clustered in a single region of the 66-kDa RT molecule. With only few exceptions, mutations that affect the RNase H activity also cause a substantial decrease in the DNA polymerase function. This suggests that, unlike the RT from murine leukemia virus (MuLV), it is difficult to genetically separate the catalytic domains responsible for the RNase H and DNA polymerase functions of HIV-1 RT. Those few mutations that differentially affect the RNase H and the polymerase activities of HIV-1 RT suggest that, as in MuLV, the polymerase domain is in the amino-terminus and the RNase H domain is in the carboxy-terminus. We have also generated chimeric molecules that are composed of sequences from the RT of HIV-1 and MuLV and these hybrid RTs were analyzed for their enzymatic properties. Two of these chimeric RTs possess RNase H activity but lack detectable DNA polymerase activity.

Original languageEnglish
Pages (from-to)575-580
Number of pages6
Issue number2
StatePublished - Apr 1990


FundersFunder number
U.S. Department of Health and Human ServicesNO1 CO-741 01
National Cancer Institute
United States-Israel Binational Science Foundation


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