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Title: Ninety-Nine Is Not Enough: Molecular Characterization of Inhibitor-Resistant Human Immunodeficiency Virus Type 1 Protease Mutants with Insertions in the Flap Region

Abstract

While the selection of amino acid insertions in human immunodeficiency virus (HIV) reverse transcriptase (RT) is a known mechanism of resistance against RT inhibitors, very few reports on the selection of insertions in the protease (PR) coding region have been published. It is still unclear whether these insertions impact protease inhibitor (PI) resistance and/or viral replication capacity. We show that the prevalence of insertions, especially between amino acids 30 to 41 of HIV type 1 (HIV-1) PR, has increased in recent years. We identified amino acid insertions at positions 33 and 35 of the PR of HIV-1-infected patients who had undergone prolonged treatment with PIs, and we characterized the contribution of these insertions to viral resistance. We prepared the corresponding mutated, recombinant PR variants with or without insertions at positions 33 and 35 and characterized them in terms of enzyme kinetics and crystal structures. We also engineered the corresponding recombinant viruses and analyzed the PR susceptibility and replication capacity by recombinant virus assay. Both in vitro methods confirmed that the amino acid insertions at positions 33 and 35 contribute to the viral resistance to most of the tested PIs. The structural analysis revealed local structural rearrangements in the flap regionmore » and in the substrate binding pockets. The enlargement of the PR substrate binding site together with impaired flap dynamics could account for the weaker inhibitor binding by the insertion mutants. Amino acid insertions in the vicinity of the binding cleft therefore represent a novel mechanism of HIV resistance development.« less

Authors:
; ; ; ; ; ; ; ; ;  [1]
  1. Quest
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
USDOE
OSTI Identifier:
1006665
Resource Type:
Journal Article
Journal Name:
J. Virol.
Additional Journal Information:
Journal Volume: 82; Journal Issue: (12) ; 06, 2008; Journal ID: ISSN 0022-538X
Country of Publication:
United States
Language:
ENGLISH
Subject:
36 MATERIALS SCIENCE; AIDS VIRUS; AMINO ACIDS; CAPACITY; CRYSTAL STRUCTURE; DRUGS; DYNAMICS; ENZYMES; INHIBITION; IN VITRO; KINETICS; MUTANTS; PATIENTS; SUBSTRATES; TRANSCRIPTION; VIRUSES

Citation Formats

Koiek, Milan, Saskova, Klara Grantz, Rezaova, Pavlina, Brynda, Jii, van Maarseveen, Noortje M, De Jong, Dorien, Boucher, Charles A, Kagan, Ron M, Nijhuis, Monique, Konvalinka, Jan, Charles U), and Utrecht). Ninety-Nine Is Not Enough: Molecular Characterization of Inhibitor-Resistant Human Immunodeficiency Virus Type 1 Protease Mutants with Insertions in the Flap Region. United States: N. p., 2008. Web. doi:10.1128/JVI.02325-07.
Koiek, Milan, Saskova, Klara Grantz, Rezaova, Pavlina, Brynda, Jii, van Maarseveen, Noortje M, De Jong, Dorien, Boucher, Charles A, Kagan, Ron M, Nijhuis, Monique, Konvalinka, Jan, Charles U), & Utrecht). Ninety-Nine Is Not Enough: Molecular Characterization of Inhibitor-Resistant Human Immunodeficiency Virus Type 1 Protease Mutants with Insertions in the Flap Region. United States. https://doi.org/10.1128/JVI.02325-07
Koiek, Milan, Saskova, Klara Grantz, Rezaova, Pavlina, Brynda, Jii, van Maarseveen, Noortje M, De Jong, Dorien, Boucher, Charles A, Kagan, Ron M, Nijhuis, Monique, Konvalinka, Jan, Charles U), and Utrecht). 2008. "Ninety-Nine Is Not Enough: Molecular Characterization of Inhibitor-Resistant Human Immunodeficiency Virus Type 1 Protease Mutants with Insertions in the Flap Region". United States. https://doi.org/10.1128/JVI.02325-07.
@article{osti_1006665,
title = {Ninety-Nine Is Not Enough: Molecular Characterization of Inhibitor-Resistant Human Immunodeficiency Virus Type 1 Protease Mutants with Insertions in the Flap Region},
author = {Koiek, Milan and Saskova, Klara Grantz and Rezaova, Pavlina and Brynda, Jii and van Maarseveen, Noortje M and De Jong, Dorien and Boucher, Charles A and Kagan, Ron M and Nijhuis, Monique and Konvalinka, Jan and Charles U) and Utrecht)},
abstractNote = {While the selection of amino acid insertions in human immunodeficiency virus (HIV) reverse transcriptase (RT) is a known mechanism of resistance against RT inhibitors, very few reports on the selection of insertions in the protease (PR) coding region have been published. It is still unclear whether these insertions impact protease inhibitor (PI) resistance and/or viral replication capacity. We show that the prevalence of insertions, especially between amino acids 30 to 41 of HIV type 1 (HIV-1) PR, has increased in recent years. We identified amino acid insertions at positions 33 and 35 of the PR of HIV-1-infected patients who had undergone prolonged treatment with PIs, and we characterized the contribution of these insertions to viral resistance. We prepared the corresponding mutated, recombinant PR variants with or without insertions at positions 33 and 35 and characterized them in terms of enzyme kinetics and crystal structures. We also engineered the corresponding recombinant viruses and analyzed the PR susceptibility and replication capacity by recombinant virus assay. Both in vitro methods confirmed that the amino acid insertions at positions 33 and 35 contribute to the viral resistance to most of the tested PIs. The structural analysis revealed local structural rearrangements in the flap region and in the substrate binding pockets. The enlargement of the PR substrate binding site together with impaired flap dynamics could account for the weaker inhibitor binding by the insertion mutants. Amino acid insertions in the vicinity of the binding cleft therefore represent a novel mechanism of HIV resistance development.},
doi = {10.1128/JVI.02325-07},
url = {https://www.osti.gov/biblio/1006665}, journal = {J. Virol.},
issn = {0022-538X},
number = (12) ; 06, 2008,
volume = 82,
place = {United States},
year = {Mon Jul 21 00:00:00 EDT 2008},
month = {Mon Jul 21 00:00:00 EDT 2008}
}