Learn More
Initiation of RNA-dependent DNA synthesis by retroviral reverse transcriptases is generally considered as unspecific. In the case of human immunodeficiency virus type 1 (HIV-1), the natural primer is tRNA3Lys. We recently found evidence of complex interactions between tRNA3Lys and HIV-1 RNA that may be involved in the priming process. In this study, we(More)
Initiation of human immunodeficiency virus type 1 (HIV-1) reverse transcription requires specific recognition of the viral genome, tRNA3Lys, which acts as primer, and reverse transcriptase (RT). The specificity of this ternary complex is mediated by intricate interactions between HIV-1 RNA and tRNA3Lys, but remains poorly understood at the three-dimensional(More)
We recently showed that primer tRNA3Lys, human immunodeficiency virus type 1 (HIV-1) RNA and HIV-1 reverse transcriptase (RT) form a specific complex of initiation of reverse transcription that can be functionally distinguished from the elongation complex, which can be obtained by substituting an 18mer oligodeoxyribonucleotide (ODN) for the natural primer(More)
We have modified an Escherichia coli vector expressing 66-kDa HIV-1 reverse transcriptase (p66) so that it simultaneously expresses this and the pol-coded protease. The twin expression cassette yields high quantities of both reverse transcriptase and protease; however, under these conditions, 50% of the over-expressed p66 reverse transcriptase is processed,(More)
Conversion of the single-stranded RNA of an invading retrovirus into double-stranded proviral DNA is catalyzed in a multi-step process by a single virus-coded enzyme, reverse transcriptase (RT). Achieving this requires a combination of DNA polymerase abd ribonuclease H (RNase H) activities, which are located at the amino and carboxy terminus of the enzyme,(More)
A contribution of the 51-kDa subunit of human immunodeficiency virus type-1 reverse transcriptase to activities of the parental heterodimer (p66/p51) was assessed in "selectively deleted" heterodimers whose p51 component contained C-terminal truncations of 13, 19, or 25 residues. Analyses included (i) efficiency of reconstitution into heterodimer, (ii)(More)
We have analysed the mechanism of ribonuclease H (RNaseH) induced cleavage of a defined RNA-DNA hybrid by human immuno-deficiency virus (HIV-1) reverse transcriptase (RT). An in vitro transcribed RNA labelled at the 3' end was hybridized to a pentadecameric DNA oligonucleotide complementary to an internal region of the RNA. Upon incubation of this RNA-DNA(More)
Human immunodeficiency virus (HIV) DNA synthesis is accompanied by degradation of genomic RNA by the RNase H of reverse transcriptase (RT). Two different modes of RNase H activity appear necessary for complete RNA removal. In one, occurring during minus strand synthesis, positioning of the RNase H is determined by binding of the polymerase active site to(More)
Recombinant proteins containing a short stretch of contiguous histidine residues (approximately 6) ("a His-tag") can be specifically bound to N-nitrilotriacetic-acid-chelated nickel ions, providing a convenient general method for their purification. A lipid derivatized with a nickel-chelating head group may provide a general approach to two-dimensional(More)