Characterization of the p68/p58 Heterodimer of Human Immunodeficiency Virus Type 2 Reverse Transcriptase

Fan, Naisheng; Rank, Kenneth B.; Poppe, Susan M.; Tarpley, W. G.; Sharma, Satish Kumar

doi:10.1021/bi9516440

Cited by 20 publications

(19 citation statements)

References 26 publications

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“…In each of our model assays, we found relatively small quantitative differences between HIV-1 and HIV-2 RNase H activities under standard assay conditions, in agreement with Fan et al (16). However, with lower enzyme concentrations, we observed that HIV-2 RT is compromised in its ability to catalyze total RNase H cleavage (Fig.…”

Section: Discussionsupporting

confidence: 91%

“…In earlier work (16,28,29,61), the DNA polymerase activity of HIV-2 RT was found to be similar to that of HIV-1 RT. The results for RNA-dependent synthesis of minus-strand DNA with an RT/primer-template ratio of 1 showed no difference in HIV-1 and HIV-2 RT activities, in agreement with these previous studies; however, when this ratio was reduced to 0.1, HIV-1 RT exhibited a higher rate and greater extent of DNA synthesis (almost 3-fold and ϳ2-fold, respectively) than the HIV-2 enzyme (Fig.…”

Section: Discussionmentioning

confidence: 83%

“…Like HIV-1 RT, the HIV-2 enzyme was shown to be a heterodimeric protein (12,15,39,44) with a large (68-kDa) subunit and a small (54-kDa) subunit generated by cleavage of p68 at Met 484 (15). In early studies, performed primarily with homopolymeric substrates (16,28,29,39,44), it was established that HIV-2 RT behaves like a true retroviral RT, having both RNA-and DNA-dependent DNA polymerase activities, as well as RNase H activity.…”

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confidence: 99%

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Human Immunodeficiency Virus Type 2 Reverse Transcriptase Activity in Model Systems That Mimic Steps in Reverse Transcription

Post

Guo

Howard

et al. 2003

J Virol

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Human immunodeficiency virus type 2 (HIV-2) infection is a serious problem in West Africa and Asia.However, there have been relatively few studies of HIV-2 reverse transcriptase (RT), a potential target for antiviral therapy. Detailed knowledge of HIV-2 RT activities is critical for development of specific highthroughput screening assays of potential inhibitors. Here, we have conducted a systematic evaluation of HIV-2 RT function, using assays that model specific steps in reverse transcription. Parallel studies were performed with HIV-1 RT. In general, under standard assay conditions, the polymerase and RNase H activities of the two enzymes were comparable. However, when the RT concentration was significantly reduced, HIV-2 RT was less active than the HIV-1 enzyme. HIV-2 RT was also impaired in its ability to catalyze secondary RNase H cleavage in assays that mimic tRNA primer removal during plus-strand transfer and degradation of genomic RNA fragments during minus-strand DNA synthesis. In addition, initiation of plus-strand DNA synthesis was much less efficient with HIV-2 RT than with HIV-1 RT. This may reflect architectural differences in the primer grip regions in the p66 (HIV-1) and p68 (HIV-2) palm subdomains of the two enzymes. The implications of our findings for antiviral therapy are discussed.

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Section: Discussionsupporting

confidence: 91%

Section: Discussionmentioning

confidence: 83%

mentioning

confidence: 99%

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Human Immunodeficiency Virus Type 2 Reverse Transcriptase Activity in Model Systems That Mimic Steps in Reverse Transcription

Post

Guo

Howard

et al. 2003

J Virol

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“…RT from 98CN009 was the least efficient enzyme in the panel for all three activities. Consistent with previous reports (14,27,49,56), the DNA polymerase and RNase H activities of RT from the HIV-2 strain EHO-287 were significantly less than those of most of the HIV-1 RTs.…”

Section: Methodssupporting

confidence: 92%

“…(Pan troglodytes troglodytes) and the 89% amino acid identity between RTs from HIV-2 group A and sooty mangabey SIV illustrate the more proximal relationships between the two HIV types and the SIV taxa from which they originated. The HIV-1 enzymes are, in general, slightly more active than their HIV-2 counterparts (14,27,49,56), which has been proposed to contribute to the greater virulence of the HIV-1 strains than of the HIV-2 strains (19,44). Many of the same factors that drive rapid viral evolution and diversification of the M group (7,8,17,40,43) have also contributed to the rapid appearance of drug resistance mutations in the RT and protease (PR) genes.…”

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confidence: 99%

Cross-Clade Inhibition of Recombinant Human Immunodeficiency Virus Type 1 (HIV-1), HIV-2, and Simian Immunodeficiency Virus SIVcpz Reverse Transcriptases by RNA Pseudoknot Aptamers

Held

Kissel

Thacker

et al. 2007

J Virol

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Reverse transcriptase (RT) remains a primary target in therapies directed at human immunodeficiency virus type 1 (HIV-1). RNA aptamers that bind RT from HIV-1 subtype B have been shown to protect human cells from infection and to reduce viral infectivity, but little is known about the sensitivity of the inhibition to amino sequence variations of the RT target. Therefore, we assembled a panel of 10 recombinant RTs from phylogenetically diverse lentiviral isolates (including strains of HIV-1, simian immunodeficiency virus SIVcpz, and HIV-2). After validating the panel by measuring enzymatic activities and inhibition by small-molecule drugs, dose-response curves for each enzyme were established for four pseudoknot RNA aptamers representing two structural subfamilies. All four aptamers potently inhibited RTs from multiple HIV-1 subtypes. For aptamers carrying family 1 pseudoknots, natural resistance was essentially all-or-none and correlated with the identity of the amino acid at position 277. In contrast, natural resistance to aptamers carrying the family 2 pseudoknots was much more heterogeneous, both in degree (gradation of 50% inhibitory concentrations) and in distribution across clades. Site-directed and subunit-specific mutagenesis identified a common R/K polymorphism within the p66 subunit as a primary determinant of resistance to family 1, but not family 2, pseudoknot aptamers. RNA structural diversity therefore translates into a nonoverlapping spectrum of mutations that confer resistance, likely due to differences in atomic-level contacts with RT.The reverse transcriptases (RTs) of the human and simian immunodeficiency viruses (HIVs and SIVs, respectively) are encoded by the viral pol gene and are expressed from viral mRNA as part of the multifunctional Gag-Pol polyprotein. Mature RT is the product of proteolytic processing of the polyprotein, first into an asymmetric homodimer and then into the mature heterodimer (21). Due to its central role in HIV type 1 (HIV-1) replication and the early clinical availability of anti-RT compounds, RT has long been an established therapeutic target. Of the 22 anti-HIV compounds currently approved by the U.S. Food and Drug Administration, 15 target the viral RT (60). The nucleoside analogue RT inhibitors (NRTIs) are deoxynucleoside triphosphate analogues that result in chain termination when incorporated by RT into a growing DNA strand. The nonnucleoside RT inhibitors (NNRTIs) bind RT near the active site and disrupt enzymatic activity by allosteric inhibition (38, 59). The search for new anti-RT compounds is fueled by cytotoxicity and clinically selected resistance-resulting from drug exclusion or from postincorporation excision (45)-which are persistent problems for both classes of RT inhibitors.Nucleic acid aptamers, ribozymes, antisense RNA, and small interfering RNA exhibit potent antiviral effects and are under development as potential gene therapy adjuvants to smallmolecule therapeutics (25,34), and several of these have recently entered into clinical trials (1,39,4...

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The ribonuclease H activity of the reverse transcriptases of human immunodeficiency viruses type 1 and type 2 is affected by the thumb subdomain of the small protein subunits11Edited by J. Karn

Sevilya

Loya

Hughes

et al. 2001

Journal of Molecular Biology

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Characterization of the p68/p58 Heterodimer of Human Immunodeficiency Virus Type 2 Reverse Transcriptase

Cited by 20 publications

References 26 publications

Human Immunodeficiency Virus Type 2 Reverse Transcriptase Activity in Model Systems That Mimic Steps in Reverse Transcription

Human Immunodeficiency Virus Type 2 Reverse Transcriptase Activity in Model Systems That Mimic Steps in Reverse Transcription

Cross-Clade Inhibition of Recombinant Human Immunodeficiency Virus Type 1 (HIV-1), HIV-2, and Simian Immunodeficiency Virus SIVcpz Reverse Transcriptases by RNA Pseudoknot Aptamers

The ribonuclease H activity of the reverse transcriptases of human immunodeficiency viruses type 1 and type 2 is affected by the thumb subdomain of the small protein subunits11Edited by J. Karn

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