Antiviral activity of recombinant ankyrin targeted to the capsid domain of HIV-1 Gag polyprotein

ABSTRACT Particle maturation is a critical step in the HIV-1 replication cycle that requires proteolytic cleavage of the Gag polyprotein into its constitutive proteins: the matrix (MA), capsid (CA), nucleocapsid (NC), and p6 proteins. The accurate and efficient cleavage of Gag is essential for virion infectivity; inhibitors of the viral protease are potent antivirals, and substitutions in Gag that prevent its cleavage result in reduced HIV-1 infectivity. In a previous study, a mutation inhibiting cleavage at the MA-CA junction was observed to potently inhibit virus infection: incorporation of small amounts of uncleaved MA-CA protein into HIV-1 particles inhibited infectivity by ∼95%, and the resulting viral particles exhibited aberrant capsids. Here we report a detailed mechanistic analysis of HIV-1 particles bearing uncleaved MA-CA protein. We show that the particles contain stable cores and can efficiently saturate host restriction by TRIMCyp in target cells. We further show that MA-CA associates with CA in particles without detectably affecting the formation of intermolecular CA interfaces. Incorporation of MA-CA did not markedly affect reverse transcription in infected cells, but nuclear entry was impaired and integration targeting was altered. Additionally, results from mutational analysis of Gag revealed that membrane-binding elements of MA contribute to the antiviral activity of uncleaved MA-CA protein. Our results suggest that small amounts of partially processed Gag subunits coassemble with CA during virion maturation, resulting in impaired capsid functions. IMPORTANCE To become infectious, newly formed HIV-1 particles undergo a process of maturation in which the viral polyproteins are cleaved into smaller components. A previous study demonstrated that inclusion of even small quantities of an uncleavable mutant Gag polyprotein results in a strong reduction in virus infectivity. Here we show that the mechanism of transdominant inhibition by uncleavable Gag involves inhibition of nuclear entry and alteration of viral integration sites. Additionally, the results of mutational analysis suggest that the membrane-binding activity of Gag is a major requirement for the antiviral activity. These results further define the antiviral mechanism of uncleavable Gag, which may be useful for exploiting this effect to develop new antivirals.

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SUMOylation of SAMHD1 at Lysine 595 is required for HIV-1 restriction in non-cycling cells

Nature Communications ◽

10.1038/s41467-021-24802-5 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Charlotte Martinat ◽

Arthur Cormier ◽

Joëlle Tobaly-Tapiero ◽

Noé Palmic ◽

Nicoletta Casartelli ◽

...

Keyword(s):

Antiviral Activity ◽

Reverse Transcription ◽

Immune Cells ◽

Viral Restriction ◽

Antiviral Function ◽

Hiv 1 ◽

Constitutively Active

AbstractSAMHD1 is a cellular triphosphohydrolase (dNTPase) proposed to inhibit HIV-1 reverse transcription in non-cycling immune cells by limiting the supply of the dNTP substrates. Yet, phosphorylation of T592 downregulates SAMHD1 antiviral activity, but not its dNTPase function, implying that additional mechanisms contribute to viral restriction. Here, we show that SAMHD1 is SUMOylated on residue K595, a modification that relies on the presence of a proximal SUMO-interacting motif (SIM). Loss of K595 SUMOylation suppresses the restriction activity of SAMHD1, even in the context of the constitutively active phospho-ablative T592A mutant but has no impact on dNTP depletion. Conversely, the artificial fusion of SUMO2 to a non-SUMOylatable inactive SAMHD1 variant restores its antiviral function, a phenotype that is reversed by the phosphomimetic T592E mutation. Collectively, our observations clearly establish that lack of T592 phosphorylation cannot fully account for the restriction activity of SAMHD1. We find that SUMOylation of K595 is required to stimulate a dNTPase-independent antiviral activity in non-cycling immune cells, an effect that is antagonized by cyclin/CDK-dependent phosphorylation of T592 in cycling cells.

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Design, Synthesis, and antiviral activity of a series of CD4-mimetic small-molecule HIV-1 entry inhibitors

Bioorganic & Medicinal Chemistry ◽

10.1016/j.bmc.2021.116000 ◽

2021 ◽

pp. 116000

Author(s):

Francesca Curreli ◽

Shahad Ahmed ◽

Sofia M. Benedict Victor ◽

Ildar R. Iusupov ◽

Evgeny A. Spiridonov ◽

...

Keyword(s):

Antiviral Activity ◽

Small Molecule ◽

Design Synthesis ◽

Entry Inhibitors ◽

Hiv 1

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Structure, Function, and Interactions of the HIV-1 Capsid Protein

Life ◽

10.3390/life11020100 ◽

2021 ◽

Vol 11 (2) ◽

pp. 100

Author(s):

Eric Rossi ◽

Megan E. Meuser ◽

Camille J. Cunanan ◽

Simon Cocklin

Keyword(s):

Life Cycle ◽

Capsid Protein ◽

Adaptor Proteins ◽

Restriction Factors ◽

Gag Polyprotein ◽

Immunodeficiency Virus ◽

Host Cell Factors ◽

Hiv 1

The capsid (CA) protein of the human immunodeficiency virus type 1 (HIV-1) is an essential structural component of a virion and facilitates many crucial life cycle steps through interactions with host cell factors. Capsid shields the reverse transcription complex from restriction factors while it enables trafficking to the nucleus by hijacking various adaptor proteins, such as FEZ1 and BICD2. In addition, the capsid facilitates the import and localization of the viral complex in the nucleus through interaction with NUP153, NUP358, TNPO3, and CPSF-6. In the later stages of the HIV-1 life cycle, CA plays an essential role in the maturation step as a constituent of the Gag polyprotein. In the final phase of maturation, Gag is cleaved, and CA is released, allowing for the assembly of CA into a fullerene cone, known as the capsid core. The fullerene cone consists of ~250 CA hexamers and 12 CA pentamers and encloses the viral genome and other essential viral proteins for the next round of infection. As research continues to elucidate the role of CA in the HIV-1 life cycle and the importance of the capsid protein becomes more apparent, CA displays potential as a therapeutic target for the development of HIV-1 inhibitors.

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Flavonol 7-O-Glucoside Herbacitrin Inhibits HIV-1 Replication through Simultaneous Integrase and Reverse Transcriptase Inhibition

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2019/1064793 ◽

2019 ◽

Vol 2019 ◽

pp. 1-6 ◽

Cited By ~ 3

Author(s):

Éva Áy ◽

Attila Hunyadi ◽

Mária Mezei ◽

János Minárovits ◽

Judit Hohmann

Keyword(s):

Antiviral Activity ◽

Reverse Transcriptase ◽

Cell Cultures ◽

Core Protein ◽

Integrase Inhibitor ◽

Host Cells ◽

Cytotoxic Activities ◽

Protein Levels ◽

Hiv 1 ◽

Antiretroviral Activity

Here we report the evaluation of the antiretroviral effect of two flavonoid 7-O-glucosides, herbacitrin (1) and gossypitrin (2), together with quercetin (3), a well-studied flavonol. Antiviral activity of the flavonoids was assessed by analyzing HIV-1 p24 core protein levels in the supernatants of HIV-1 infected MT-4 and MT-2 cell cultures. The compounds showed mild to weak cytotoxic activities on the host cells; herbacitrin was the strongest in this regard (CC50=27.8 and 63.64 μM on MT-4 and MT-2 cells, respectively). In nontoxic concentrations, herbacitrin and quercetin reduced HIV-1 replication, whereas gossypitrin was ineffective. Herbacitrin was found to inhibit reverse transcriptase at 21.5 μM, while it was a more potent integrase inhibitor already active at 2.15 μM. Therefore, our observations suggest that herbacitrin exerts antiretroviral activity through simultaneously acting on these two targets of HIV-1 and that integrase inhibition might play a major role in this activity.

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Phenotypic Susceptibility to Bevirimat in Isolates from HIV-1-Infected Patients without Prior Exposure to Bevirimat

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01784-09 ◽

2010 ◽

Vol 54 (6) ◽

pp. 2345-2353 ◽

Cited By ~ 39

Author(s):

Nicolas A. Margot ◽

Craig S. Gibbs ◽

Michael D. Miller

Keyword(s):

Recombinant Viruses ◽

Gag Polyprotein ◽

New Class ◽

Major Mutation ◽

Hiv Drugs ◽

The Impact ◽

First Time ◽

Hiv 1

ABSTRACT Bevirimat (BVM) is the first of a new class of anti-HIV drugs with a novel mode of action known as maturation inhibitors. BVM inhibits the last cleavage of the Gag polyprotein by HIV-1 protease, leading to the accumulation of the p25 capsid-small peptide 1 (SP1) intermediate and resulting in noninfectious HIV-1 virions. Early clinical studies of BVM showed that over 50% of the patients treated with BVM did not respond to treatment. We investigated the impact of prior antiretroviral (ARV) treatment and/or natural genetic diversity on BVM susceptibility by conducting in vitro phenotypic analyses of viruses made from patient samples. We generated 31 recombinant viruses containing the entire gag and protease genes from 31 plasma samples from HIV-1-infected patients with (n = 21) or without (n = 10) prior ARV experience. We found that 58% of the patient isolates tested had a >10-fold reduced susceptibility to BVM, regardless of the patient's ARV experience or the level of isolate resistance to protease inhibitors. Analysis of mutants with site-directed mutations confirmed the role of the V370A SP1 polymorphism (SP1-V7A) in resistance to BVM. Furthermore, we demonstrated for the first time that a capsid polymorphism, V362I (CA protein-V230I), is also a major mutation conferring resistance to BVM. In contrast, none of the previously defined resistance-conferring mutations in Gag selected in vitro (H358Y, L363M, L363F, A364V, A366V, or A366T) were found to occur among the viruses that we analyzed. Our results should be helpful in the design of diagnostics for prediction of the potential benefit of BVM treatment in HIV-1-infected patients.

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Antiviral activity of derivatized dextrans on HIV-1 infection of primary macrophages and blood lymphocytes

Biochimica et Biophysica Acta (BBA) - Molecular Basis of Disease ◽

10.1016/s0925-4439(97)00057-4 ◽

1997 ◽

Vol 1362 (1) ◽

pp. 47-55 ◽

Cited By ~ 4

Author(s):

Nabila Seddiki ◽

Elisabeth Mbemba ◽

Didier Letourneur ◽

Loyda Ylisastigui ◽

Abdelaziz Benjouad ◽

...

Keyword(s):

Antiviral Activity ◽

Blood Lymphocytes ◽

Hiv 1

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Towards Inhibition of Vif-APOBEC3G Interaction: Which Protein to Target?

Advances in Virology ◽

10.1155/2010/649315 ◽

2010 ◽

Vol 2010 ◽

pp. 1-10 ◽

Cited By ~ 4

Author(s):

Iris Cadima-Couto ◽

Joao Goncalves

Keyword(s):

Antiviral Activity ◽

Protein Binding ◽

Mechanism Of Action ◽

Cellular Immunity ◽

Viral Infectivity ◽

Binding Partners ◽

Viral Infectivity Factor ◽

Vif Protein ◽

Hiv 1 ◽

Antiretroviral Activity

APOBEC proteins appeared in the cellular battle against HIV-1 as part of intrinsic cellular immunity. The antiretroviral activity of some of these proteins is overtaken by the action of HIV-1 Viral Infectivity Factor (Vif) protein. Since the discovery of APOBEC3G (A3G) as an antiviral factor, many advances have been made to understand its mechanism of action in the cell and how Vif acts in order to counteract its activity. The mainstream concept is that Vif overcomes the innate antiviral activity of A3G by direct protein binding and promoting its degradation via the cellular ubiquitin/proteasomal pathway. Vif may also inhibit A3G through mechanisms independent of proteasomal degradation. Binding of Vif to A3G is essential for its degradation since disruption of this interaction is predicted to stimulate intracellular antiviral immunity. In this paper we will discuss the different binding partners between both proteins as one of the major challenges for the development of new antiviral drugs.

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Correction: Normalization strategy for the LC-MS bioanalysis of protein kinetics assays via internal proteolytic analyte utilized as control standard: application in studies of HIV-1 protease cleavage of HIV-1 Gag polyprotein in HIV maturation inhibition research

Analytical Methods ◽

10.1039/c7ay90125a ◽

2017 ◽

Vol 9 (37) ◽

pp. 5557-5557

Author(s):

Joseph L. Cantone ◽

Zeyu Lin ◽

Ira B. Dicker ◽

Dieter M. Drexler

Keyword(s):

Protein Kinetics ◽

Gag Polyprotein ◽

Protease Cleavage ◽

Normalization Strategy ◽

Standard Application ◽

Hiv 1

Correction for ‘Normalization strategy for the LC-MS bioanalysis of protein kinetics assays via internal proteolytic analyte utilized as control standard: application in studies of HIV-1 protease cleavage of HIV-1 Gag polyprotein in HIV maturation inhibition research’ by Joseph L. Cantone et al., Anal. Methods, 2017, DOI: 10.1039/c7ay01666b.

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In Vitro Antiviral Activity and Cross-Resistance Profile of PL-100, a Novel Protease Inhibitor of Human Immunodeficiency Virus Type 1

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00149-07 ◽

2007 ◽

Vol 51 (11) ◽

pp. 4036-4043 ◽

Cited By ~ 21

Author(s):

Serge Dandache ◽

Guy Sévigny ◽

Jocelyn Yelle ◽

Brent R. Stranix ◽

Neil Parkin ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Antiviral Activity ◽

Highly Active Antiretroviral Therapy ◽

Cross Resistance ◽

Drug Resistant ◽

Resistance Profile ◽

Immunodeficiency Virus ◽

Hiv 1

ABSTRACT Despite the success of highly active antiretroviral therapy, the current emergence and spread of drug-resistant variants of human immunodeficiency virus (HIV) stress the need for new inhibitors with distinct properties. We designed, produced, and screened a library of compounds based on an original l-lysine scaffold for their potentials as HIV type 1 (HIV-1) protease inhibitors (PI). One candidate compound, PL-100, emerged as a specific and noncytotoxic PI that exhibited potent inhibition of HIV-1 protease and viral replication in vitro (Ki , ∼36 pM, and 50% effective concentration [EC50], ∼16 nM, respectively). To confirm that PL-100 possessed a favorable resistance profile, we performed a cross-resistance study using a panel of 63 viral strains from PI-experienced patients selected for the presence of primary PI mutations known to confer resistance to multiple PIs now in clinical use. The results showed that PL-100 retained excellent antiviral activity against almost all of these PI-resistant viruses and that its performance in this regard was superior to those of atazanavir, amprenavir, indinavir, lopinavir, nelfinavir, and saquinavir. In almost every case, the increase in the EC50 for PL-100 observed with viruses containing multiple mutations in protease was far less than that obtained with the other drugs tested. These data underscore the potential for PL-100 to be used in the treatment of drug-resistant HIV disease and argue for its further development.

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