Purine analog substitution of the HIV-1 polypurine tract primer defines regions controlling initiation of plus-strand DNA synthesis

Rausch, Jason W.; Grice, Stuart F. J. Le

doi:10.1093/nar/gkl909

Cited by 21 publications

(18 citation statements)

References 47 publications

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“…Despite extensive mutagenesis analysis (Rausch and Le Grice 2007), the structural basis for resistance of PPT-containing RNA/DNA hybrids to RNase H-mediated cleavage remains unclear. However, structural studies that combined mass spectrometry with NMR , raise the possibility that the PPT could be targeted by small molecules designed to antagonize its selection or removal from nascent (+) DNA.…”

Section: Protein Synthesis and Ribosomal Frameshifting: A Delicate Bamentioning

confidence: 99%

Targeting the HIV RNA Genome: High-Hanging Fruit Only Needs a Longer Ladder

Grice

2015

Current Topics in Microbiology and Immunology

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Small molecules targeting the enzymes responsible for human immunodeficiency virus (HIV) maturation, DNA synthesis and its subsequent chromosomal integration as ribonucleotide-free double-stranded DNA remain the mainstay of combination antiretroviral therapy. For infected individuals harboring drug-susceptible virus, this approach has afforded complete or near-complete viral suppression. However, in the absence of a curative strategy, the predictable emergence of drug-resistant variants requires continued development of improved antiviral strategies, inherent to which is the necessity of identifying novel targets. Regulatory elementsRegulatory elements that mediate transcription, translation, nucleocytoplasmic transport, dimerization, packaging and reverse transcription of the (+) strand RNA genomeRNA genome should now be considered viable targets for small molecule, peptide- and oligonucleotide-based therapeuticsTherapeutics . Where target specificity and cellular penetration and toxicity have been the primary obstacle to successful "macromolecule therapeutics", this chapter summarizes (a) novel approaches targeting RNA motifs whose three-dimensional structure is critical for biological function and consequently may be less prone to resistance-conferring mutations and (b) improved methods for deliveryDelivery .

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Section: Protein Synthesis and Ribosomal Frameshifting: A Delicate Bamentioning

confidence: 99%

Targeting the HIV RNA Genome: High-Hanging Fruit Only Needs a Longer Ladder

Grice

2015

Current Topics in Microbiology and Immunology

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“…There are no base contacts with positions −4 or −7, but there are phosphate contacts from −4 to −9 in the DNA strand with the RNase H primer grip region. Using model substrates and purified enzyme, positions −2 and −4 are essential for correct cleavage of the PPT (Pullen et al, 1993) and substitutions at positions +1, −2, and −4 of the PPT substantially decrease generation of the PPT primer (Rausch and Le Grice, 2007). In other studies examining the recognition and cleavage of the PPT by HIV-1 RNase H, the effects of mutations in the PPT and in the primer grip region of the RNase H domain have been assessed both in vitro and in vivo Julias et al, 2004;McWilliams et al, 2006;Jones and Hughes, 2007).…”

Section: Generation Of Ppt Primermentioning

confidence: 99%

RNase H activity: Structure, specificity, and function in reverse transcription

2008

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This review compares the well-studied RNase H activities of human immunodeficiency virus, type 1 (HIV-1) and Moloney murine leukemia virus (MoMLV) reverse transcriptases. The RNase H domains of HIV-1 and MoMLV are structurally very similar, with functions assigned to conserved subregions like the RNase H primer grip and the connection subdomain, as well as to distinct features like the C-helix and loop in MoMLV RNase H. Like cellular RNases H, catalysis by the retroviral enzymes appears to involve a two-metal ion mechanism. Unlike cellular RNases H, the retroviral RNases H display three different modes of cleavage: internal, DNA 3′ end-directed, and RNA 5′ enddirected. All three modes of cleavage appear to have roles in reverse transcription. Nucleotide sequence is an important determinant of cleavage specificity with both enzymes exhibiting a preference for specific nucleotides at discrete positions flanking an internal cleavage site as well as during tRNA primer removal and plus-strand primer generation. RNA 5′ end-directed and DNA 3′ end-directed cleavages show similar sequence preferences at the positions closest to a cleavage site. A model for how RNase H selects cleavage sites is presented that incorporates both sequence preferences and the concept of a defined window for allowable cleavage from a recessed end. Finally, the RNase H activity of HIV-1 is considered as a target for anti-virals as well as a participant in drug resistance. KeywordsRNase H; reverse transcriptase; human immunodeficiency virus; type 1 (HIV -1); Moloney murine leukemia virus (MoMLV); reverse transcription; polypurine tract (PPT)

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“…A variety of studies have identified the nucleotide positions within the PPT that are critical for proper cleavage and although some of these overlap with the more general preferences for internal cleavage, other positions do not. Thus, for proper PPT primer generation by M‐MLV RNase H, positions −1, −2, −4, −5, −6, −7, −10 and −11 are important [51,94,95], whereas positions +1, −2, −4, −5 and −7 have been found to be critical for HIV‐1 PPT primer formation [38,40,89,96–98].…”

Section: Roles Of Rnase H In Reverse Transcriptionmentioning

confidence: 99%

Ribonuclease H: properties, substrate specificity and roles in retroviral reverse transcription

Champoux

Schultz

2009

The FEBS Journal

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At the time of its discovery in 1970, the presence of an RNA-dependent DNA polymerase activity in retrovirus particles provided strong and exciting support for the hypothesis that the single-stranded RNA genome of a retrovirus is replicated through a DNA intermediate [1,2]. Not only did this discovery of reverse transcriptase (as it was dubbed) challenge the existing dogma concerning the flow of genetic information in biology, it raised the critical question as to how the DNA ⁄ RNA hybrid created when the viral genome RNA is used as a template by reverse transcriptase is further processed. In retrospect, it is not surprising that an RNase H activity that degrades the RNA strand of a DNA ⁄ RNA hybrid is required to free the newly made DNA strand (called the minus strand because it is complementary to the plus genome RNA) for use as a template in the synthesis of the second or plus strand DNA. However, it was a surprise when the retroviral-specific RNase H activity turned out to be present in the same protein molecule as the polymerase activity [3]. This intimate association of the DNA polymerase and RNase H activities in reverse transcriptase has profound effects on the activities and capabilities of both enzymes.This minireview provides a summary of the salient features of retroviral RNases H with a focus on how the shared substrate-binding sites for the two activities of reverse transcriptase endow the retroviral RNases H with features not found in the cellular counterparts, and how these unusual properties are crucial for the multiple roles played by RNase H in reverse transcription. Although occasional reference is made to other retroviral enzymes, the primary focus is on the wellstudied RNase H activities associated with human Retroviral reverse transcriptases possess both a DNA polymerase and an RNase H activity. The linkage with the DNA polymerase activity endows the retroviral RNases H with unique properties not found in the cellular counterparts. In addition to the typical endonuclease activity on a DNA ⁄ RNA hybrid, cleavage by the retroviral enzymes is also directed by both DNA 3¢ recessed and RNA 5¢ recessed ends, and by certain nucleotide sequence preferences in the vicinity of the cleavage site. This spectrum of specificities enables retroviral RNases H to carry out a series of cleavage reactions during reverse transcription that degrade the viral RNA genome after minus-strand synthesis, precisely generate the primer for the initiation of plus strands, facilitate the initiation of plus-strand synthesis and remove both plus-and minus-strand primers after they have been extended.

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Purine analog substitution of the HIV-1 polypurine tract primer defines regions controlling initiation of plus-strand DNA synthesis

Cited by 21 publications

References 47 publications

Targeting the HIV RNA Genome: High-Hanging Fruit Only Needs a Longer Ladder

Targeting the HIV RNA Genome: High-Hanging Fruit Only Needs a Longer Ladder

RNase H activity: Structure, specificity, and function in reverse transcription

Ribonuclease H: properties, substrate specificity and roles in retroviral reverse transcription

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