Proteolytic processing of an HIV-1 pol polyprotein precursor: insights into the mechanism of reverse transcriptase p66/p51 heterodimer formation

2004 ◽  
Vol 36 (9) ◽  
pp. 1836-1847 ◽  
Author(s):  
N SLUISCREMER
Keyword(s):  
Reverse Transcriptase ◽  
Proteolytic Processing ◽  
Polyprotein Precursor ◽  
Hiv 1

scholarly journals Efavirenz enhances the proteolytic processing of an HIV-1 pol polyprotein precursor and reverse transcriptase homodimer formation

FEBS Letters ◽  
2004 ◽  
Vol 579 (2) ◽  
pp. 379-384 ◽  
Author(s):  
Gilda Tachedjian ◽  
Katie L. Moore ◽  
Stephen P. Goff ◽  
Nicolas Sluis-Cremer
Keyword(s):  
Reverse Transcriptase ◽  
Proteolytic Processing ◽  
Polyprotein Precursor ◽  
Hiv 1

scholarly journals Crystal structure of prototype foamy virus (PFV) protease-reverse transcriptase fusion (PR-RT) reveals conformational plasticity: implications for function

2020 ◽  
Author(s):  
Jerry Joe E. K. Harrison ◽  
Steve Tuske ◽  
Kalyan Das ◽  
Francesc X. Ruiz ◽  
Joseph D. Bauman ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Reverse Transcriptase ◽  
Foamy Virus ◽  
Rnase H ◽  
Proteolytic Processing ◽  
Nucleic Acid Binding ◽  
Foamy Viruses ◽  
Conformational Plasticity ◽  
Polyprotein Precursor ◽  
Hiv 1

AbstractProteolytic processing of the retroviral Pol polyprotein precursor produces protease (PR), reverse transcriptase (RT), and integrase (IN), except in foamy viruses (FVs) where only the IN domain is released. Here, we report the 2.9 Å resolution crystal structure of the mature PR-RT from prototype FV (PFV) needed for processing and reverse transcription. The monomeric PFV PR exhibits similar architecture as the HIV-1 PR but the N- and C-terminal residues are unstructured. A C-terminal extension of the PR folds into two helices that supports the RT palm subdomain and anchors the PR next to the RT. The subdomains of RT: fingers, palm, thumb, and connection, and the RNase H domain, are connected by flexible linkers and spatially arranged similarly to those in the HIV-1 RT p51 subunit. Significant spatial and conformational domain rearrangements are required for nucleic acid binding. This offers structural insight into retroviral RT conformational maturation and architecture of immature enzymes.

scholarly journals Crystal Structure of a Retroviral Polyprotein: Prototype Foamy Virus Protease-Reverse Transcriptase (PR-RT)

Viruses ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1495
Author(s):  
Jerry Joe E. K. Harrison ◽  
Steve Tuske ◽  
Kalyan Das ◽  
Francesc X. Ruiz ◽  
Joseph D. Bauman ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Reverse Transcriptase ◽  
Foamy Virus ◽  
Polymerase Activity ◽  
Rnase H ◽  
Proteolytic Processing ◽  
Nucleic Acid Binding ◽  
Foamy Viruses ◽  
Polyprotein Precursor ◽  
Hiv 1

In most cases, proteolytic processing of the retroviral Pol portion of the Gag-Pol polyprotein precursor produces protease (PR), reverse transcriptase (RT), and integrase (IN). However, foamy viruses (FVs) express Pol separately from Gag and, when Pol is processed, only the IN domain is released. Here, we report a 2.9 Å resolution crystal structure of the mature PR-RT from prototype FV (PFV) that can carry out both proteolytic processing and reverse transcription but is in a configuration not competent for proteolytic or polymerase activity. PFV PR-RT is monomeric and the architecture of PFV PR is similar to one of the subunits of HIV-1 PR, which is a dimer. There is a C-terminal extension of PFV PR (101-145) that consists of two helices which are adjacent to the base of the RT palm subdomain, and anchors PR to RT. The polymerase domain of PFV RT consists of fingers, palm, thumb, and connection subdomains whose spatial arrangements are similar to the p51 subunit of HIV-1 RT. The RNase H and polymerase domains of PFV RT are connected by flexible linkers. Significant spatial and conformational (sub)domain rearrangements are therefore required for nucleic acid binding. The structure of PFV PR-RT provides insights into the conformational maturation of retroviral Pol polyproteins.

Disruption of a Salt Bridge between Asp 488 and Lys 465 in HIV-1 Reverse Transcriptase Alters Its Proteolytic Processing and Polymerase Activity

Virology ◽  
1993 ◽  
Vol 196 (2) ◽  
pp. 731-738 ◽  
Author(s):  
Laura Goobar-Larsson ◽  
Kristina Bäckbro ◽  
Torsten Unge ◽  
Ramagauri Bhikhabhai ◽  
Lotta Vrang ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Salt Bridge ◽  
Polymerase Activity ◽  
Proteolytic Processing ◽  
Hiv 1

scholarly journals Analysis of the Contribution of Reverse Transcriptase and Integrase Proteins to Retroviral RNA Dimer Conformation

2005 ◽  
Vol 79 (10) ◽  
pp. 6338-6348 ◽  
Author(s):  
Penelope Buxton ◽  
Gilda Tachedjian ◽  
Johnson Mak
Keyword(s):  
Reverse Transcriptase ◽  
Leukemia Virus ◽  
Proteolytic Processing ◽  
Moloney Murine Leukemia Virus ◽  
Stable Conformation ◽  
Rna Dimerization ◽  
Immunodeficiency Virus ◽  
Rna Dimer ◽  
Hiv 1

ABSTRACT All retroviruses contain two copies of genomic RNA that are linked noncovalently. The dimeric RNA of human immunodeficiency virus type 1 (HIV-1) undergoes rearrangement during virion maturation, whereby the dimeric RNA genome assumes a more stable conformation. Previously, we have shown that the packaging of the HIV-1 polymerase (Pol) proteins reverse transcriptase (RT) and integrase (IN) is essential for the generation of the mature RNA dimer conformation. Analysis of HIV-1 mutants that are defective in processing of Pol showed that these mutant virions contained altered dimeric RNA conformation, indicating that the mature RNA dimer conformation in HIV-1 requires the correct proteolytic processing of Pol. The HIV-1 Pol proteins are multimeric in their mature enzymatically active forms; RT forms a heterodimer, and IN appears to form a homotetramer. Using RT and IN multimerization defective mutants, we have found that dimeric RNA from these mutant virions has the same stability and conformation as wild-type RNA dimers, showing that the mature enzymatically active RT and IN proteins are dispensable for the generation of mature RNA dimer conformation. This also indicated that formation of the mature RNA dimer structure occurs prior to RT or IN maturation. We have also investigated the requirement of Pol for RNA dimerization in both Mason-Pfizer monkey virus (M-PMV) and Moloney murine leukemia virus (MoMuLV) and found that in contrast to HIV-1, Pol is dispensable for RNA dimer maturation in M-PMV and MoMuLV, demonstrating that the requirement of Pol in retroviral RNA dimer maturation is not conserved among all retroviruses.

scholarly journals Identification of amino acid residues in HIV-1 reverse transcriptase that are critical for the proteolytic processing of Gag-Pol precursors

FEBS Letters ◽  
2011 ◽  
Vol 585 (21) ◽  
pp. 3372-3377 ◽  
Author(s):  
Hironori Nishitsuji ◽  
Masaru Yokoyama ◽  
Hironori Sato ◽  
Suguru Yamauchi ◽  
Hiroshi Takaku
Keyword(s):  
Amino Acid ◽  
Reverse Transcriptase ◽  
Proteolytic Processing ◽  
Amino Acid Residues ◽  
Hiv 1

scholarly journals Cryo-EM Structure of the Pol Polyprotein Provides Insights into HIV Maturation

2021 ◽  
Author(s):  
Jerry Joe Ebow Kingsley Harrison ◽  
Dario Oliveira Passos ◽  
Jessica F Bruhn ◽  
Joseph D Bauman ◽  
Lynda Tuberty ◽  
...  
Keyword(s):  
High Resolution ◽  
Reverse Transcriptase ◽  
Antiretroviral Drugs ◽  
Proteolytic Processing ◽  
Full Length ◽  
Viral Protease ◽  
Functional Implications ◽  
Catalytic Mechanisms ◽  
Hiv 1

Many retroviral proteins are initially translated from unspliced full-length RNA as polyprotein precursors that are subsequently processed by the viral protease (PR) to yield the mature forms. In HIV-1, the enzymes, PR, reverse transcriptase (RT), and integrase (IN), are produced as part of the Gag-Pol polyprotein. While structures of the mature proteins have aided our understanding of catalytic mechanisms and the design of antiretroviral drugs, knowledge of the architecture and functional implications of the immature forms prior to PR-mediated cleavage is limited. We developed a system to produce and purify the HIV-1 Pol polyprotein intermediate precursor and determined its high-resolution cryo-EM structure. The RT portion of the polyprotein has an architecture similar to the mature RT p66/p51 heterodimer, and dimerization of the RT portion draws together two PR monomers to activate proteolytic processing. HIV-1 thus may leverage the dimerization interfaces in Pol to regulate the assembly and maturation of the polyprotein precursors.

scholarly journals A Single Amino Acid Substitution in HIV-1 Reverse Transcriptase Significantly Reduces Virion Release

2009 ◽  
Vol 84 (2) ◽  
pp. 976-982 ◽  
Author(s):  
Chien-Cheng Chiang ◽  
Shiu-Mei Wang ◽  
Yen-Yu Pan ◽  
Kuo-Jung Huang ◽  
Chin-Tien Wang
Keyword(s):  
Amino Acid ◽  
Reverse Transcriptase ◽  
Virus Replication ◽  
Amino Acid Substitution ◽  
Proteolytic Processing ◽  
Repeat Motif ◽  
Single Amino Acid Substitution ◽  
Single Amino Acid ◽  
Cleavage Efficiency ◽  
Hiv 1

ABSTRACT HIV-1 protease (PR) mediates the proteolytic processing of virus particles during or after virus budding. PR activation is thought to be triggered by appropriate Gag-Pol/Gag-Pol interaction; factors affecting this interaction either enhance or reduce PR-mediated cleavage efficiency, resulting in markedly reduced virion production or the release of inadequately processed virions. We previously showed that a Gag-Pol deletion mutation involving the reverse transcriptase tryptophan (Trp) repeat motif markedly impairs PR-mediated virus maturation and that an alanine substitution at W401 (W401A) or at both W401 and W402 (W401A/W402A) partially or almost completely negates the enhancement effect of efavirenz (a nonnucleoside reverse transcriptase inhibitor) on PR-mediated virus processing efficiency. These data suggest that the Trp repeat motif may contribute to the PR activation process. Here we demonstrate that due to enhanced Gag cleavage efficiency, W402 alanine or leucine substitution significantly reduces virus production. However, W402 replacement with phenylalanine does not significantly affect virus particle assembly or processing, but it does markedly impair viral infectivity in a single-cycle infection assay. Our results demonstrate that a single amino acid substitution at HIV-1 RT can radically affect virus assembly by enhancing Gag cleavage efficiency, suggesting that in addition to contributing to RT biological function during the early stages of virus replication, the HIV-1 RT tryptophan repeat motif in a Gag-Pol context may play an important role in suppressing the premature activation of PR during late-stage virus replication.

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