scholarly journals The Conserved Carboxy Terminus of the Capsid Domain of Human Immunodeficiency Virus Type 1 Gag Protein Is Important for Virion Assembly and Release

2004 ◽  
Vol 78 (18) ◽  
pp. 9675-9688 ◽  
Author(s):  
Daniel Melamed ◽  
Michal Mark-Danieli ◽  
Michal Kenan-Eichler ◽  
Osnat Kraus ◽  
Asher Castiel ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Hinge Region ◽  
Virion Assembly ◽  
C Terminus ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Ca Protein

ABSTRACT The retroviral Gag precursor plays an important role in the assembly of virion particles. The capsid (CA) protein of the Gag molecule makes a major contribution to this process. In the crystal structure of the free CA protein of the human immunodeficiency virus type 1 (HIV-1), 11 residues of the C terminus were found to be unstructured, and to date no information exists on the structure of these residues in the context of the Gag precursor molecule. We performed phylogenetic analysis and demonstrated a high degree of conservation of these 11 amino acids. Deletion of this cluster or introduction of various point mutations into these residues resulted in significant impairment of particle infectivity. In this cluster, two putative structural regions were identified, residues that form a hinge region (353-VGGP-356) and those that contribute to an α-helix (357-GHKARVL-363). Overall, mutations in these regions resulted in inhibition of virion production, but mutations in the hinge region demonstrated the most significant reduction. Although all the Gag mutants appeared to have normal Gag-Gag and Gag-RNA interactions, the hinge mutants were characterized by abnormal formation of cytoplasmic Gag complexes. Gag proteins with mutations in the hinge region demonstrated normal membrane association but aberrant rod-like membrane structures. More detailed analysis of these structures in one of the mutants demonstrated abnormal trapped Gag assemblies. These data suggest that the conserved CA C terminus is important for HIV-1 virion assembly and release and define a putative target for drug design geared to inhibit the HIV-1 assembly process.

scholarly journals In Vitro Resistance to the Human Immunodeficiency Virus Type 1 Maturation Inhibitor PA-457 (Bevirimat)

2006 ◽  
Vol 80 (22) ◽  
pp. 10957-10971 ◽  
Author(s):  
Catherine S. Adamson ◽  
Sherimay D. Ablan ◽  
Ioana Boeras ◽  
Ritu Goila-Gaur ◽  
Ferri Soheilian ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Single Amino Acid ◽  
Compensatory Mutation ◽  
C Terminus ◽  
Immunodeficiency Virus ◽  
Maturation Inhibitor ◽  
Hiv 1

ABSTRACT 3-O-(3′,3′-dimethylsuccinyl)betulinic acid (PA-457 or bevirimat) potently inhibits human immunodeficiency virus type 1 (HIV-1) maturation by blocking a late step in the Gag processing pathway, specifically the cleavage of SP1 from the C terminus of capsid (CA). To gain insights into the mechanism(s) by which HIV-1 could evolve resistance to PA-457 and to evaluate the likelihood of such resistance arising in PA-457-treated patients, we sought to identify and characterize a broad spectrum of HIV-1 variants capable of conferring resistance to this compound. Numerous independent rounds of selection repeatedly identified six single-amino-acid substitutions that independently confer PA-457 resistance: three at or near the C terminus of CA (CA-H226Y, -L231F, and -L231M) and three at the first and third residues of SP1 (SP1-A1V, -A3T, and -A3V). We determined that mutations CA-H226Y, CA-L231F, CA-L231M, and SP1-A1V do not impose a significant replication defect on HIV-1 in culture. In contrast, mutations SP1-A3V and -A3T severely impaired virus replication and inhibited virion core condensation. The replication defect imposed by SP1-A3V was reversed by a second-site compensatory mutation in CA (CA-G225S). Intriguingly, high concentrations of PA-457 enhanced the maturation of SP1 residue 3 mutants. The different phenotypes associated with mutations that confer PA-457 resistance suggest the existence of multiple mechanisms by which HIV-1 can evolve resistance to this maturation inhibitor. These findings have implications for the ongoing development of PA-457 to treat HIV-1 infection in vivo.

scholarly journals Chimeric Human Immunodeficiency Virus Type 1 (HIV-1) Virions Containing HIV-2 or Simian Immunodeficiency Virus Nef Are Resistant to Cyclosporine Treatment

2004 ◽  
Vol 78 (4) ◽  
pp. 1843-1850 ◽  
Author(s):  
Mahfuz Khan ◽  
Lingling Jin ◽  
Ming Bo Huang ◽  
Lesa Miles ◽  
Vincent C. Bond ◽  
...  
Keyword(s):  
Amino Acids ◽  
Human Immunodeficiency Virus ◽  
Simian Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Cyclophilin A ◽  
C Terminus ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT The viral protein Nef and the cellular factor cyclophilin A are both required for full infectivity of human immunodeficiency virus type 1 (HIV-1) virions. In contrast, HIV-2 and simian immunodeficiency virus (SIV) do not incorporate cyclophilin A into virions or need it for full infectivity. Since Nef and cyclophilin A appear to act in similar ways on postentry events, we determined whether chimeric HIV-1 virions that contained either HIV-2 or SIV Nef would have a direct effect on cyclophilin A dependence. Our results show that chimeric HIV-1 virions containing either HIV-2 or SIV Nef are resistant to treatment by cyclosporine and enhance the infectivity of virions with mutations in the cyclophilin A binding loop of Gag. Amino acids at the C terminus of HIV-2 and SIV are necessary for inducing cyclosporine resistance. However, transferring these amino acids to the C terminus of HIV-1 Nef is insufficient to induce cyclosporine resistance in HIV-1. These results suggest that HIV-2 and SIV Nef are able to compensate for the need for cyclophilin A for full infectivity and that amino acids present at the C termini of these proteins are important for this function.

scholarly journals Particle Size Determinants in the Human Immunodeficiency Virus Type 1 Gag Protein

1998 ◽  
Vol 72 (6) ◽  
pp. 4667-4677 ◽  
Author(s):  
Laurence Garnier ◽  
Lee Ratner ◽  
Benjamin Rovinski ◽  
Shi-Xian Cao ◽  
John W. Wills
Keyword(s):  
Human Immunodeficiency Virus ◽  
Particle Size ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Gag Protein ◽  
Important Region ◽  
C Terminus ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT The retroviral Gag protein plays the central role in the assembly process and can form membrane-enclosed, virus-like particles in the absence of any other viral products. These particles are similar to authentic virions in density and size. Three small domains of the human immunodeficiency virus type 1 (HIV-1) Gag protein have been previously identified as being important for budding. Regions that lie outside these domains can be deleted without any effect on particle release or density. However, the regions of Gag that control the size of HIV-1 particles are less well understood. In the case of Rous sarcoma virus (RSV), the size determinant maps to the CA (capsid) and adjacent spacer sequences within Gag, but systematic mapping of the HIV Gag protein has not been reported. To locate the size determinants of HIV-1, we analyzed a large collection of Gag mutants. To our surprise, all mutants with defects in the MA (matrix), CA, and the N-terminal part of NC (nucleocapsid) sequences produced dense particles of normal size, suggesting that oncoviruses (RSV) and lentiviruses (HIV-1) have different size-controlling elements. The most important region found to be critical for determining HIV-1 particle size is the p6 sequence. Particles lacking all or small parts of p6 were uniform in size distribution but very large as measured by rate zonal gradients. Further evidence for this novel function of p6 was obtained by placing this sequence at the C terminus of RSV CA mutants that produce heterogeneously sized particles. We found that the RSV-p6 chimeras produced normally sized particles. Thus, we present evidence that the entire p6 sequence plays a role in determining the size of a retroviral particle.

scholarly journals Capsid Proteins from Human Immunodeficiency Virus Type 1 and Simian Immunodeficiency Virus SIVmac Can Coassemble into Mature Cores of Infectious Viruses

2008 ◽  
Vol 82 (17) ◽  
pp. 8253-8261 ◽  
Author(s):  
Jianbo Chen ◽  
Vinay K. Pathak ◽  
Weiqun Peng ◽  
Wei-Shau Hu
Keyword(s):  
Human Immunodeficiency Virus ◽  
Simian Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Infectious Virus ◽  
Virus Infectivity ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Ca Protein

ABSTRACT We have recently shown that the Gag polyproteins from human immunodeficiency virus type 1 (HIV-1) and HIV-2 can coassemble and functionally complement each other. During virion maturation, the Gag polyproteins undergo proteolytic cleavage to release mature proteins including capsid (CA), which refolds and forms the outer shell of a cone-shaped mature core. Less than one-half of the CA proteins present within the HIV-1 virion are required to form the mature core. Therefore, it is unclear whether the mature core in virions containing both HIV-1 and HIV-2 Gag consists of CA proteins from a single virus or from both viruses. To determine whether CA proteins from two different viruses can coassemble into mature cores of infectious viruses, we exploited the specificity of the tripartite motif 5α protein from the rhesus monkey (rhTRIM5α) for cores containing HIV-1 CA (hCA) but not the simian immunodeficiency virus SIVmac CA protein (sCA). If hCA and sCA cannot coassemble into the same core when equal amounts of sCA and hCA are coexpressed, the infectivities of such virus preparations in cells should be inhibited less than twofold by rhTRIM5α. However, if hCA and sCA can coassemble into the same core structure to form a mixed core, rhTRIM5α would be able to recognize such cores and significantly restrict virus infectivity. We examined the restriction phenotypes of viruses containing both hCA and sCA. Our results indicate that hCA and sCA can coassemble into the same mature core to produce infectious virus. To our knowledge, this is the first demonstration of functional coassembly of heterologous CA protein into the retroviral core.

scholarly journals Inhibition of Human Immunodeficiency Virus Type 1 Replication in Human Cells by Debio-025, a Novel Cyclophilin Binding Agent

2008 ◽  
Vol 52 (4) ◽  
pp. 1302-1317 ◽  
Author(s):  
Roger G. Ptak ◽  
Philippe A. Gallay ◽  
Dirk Jochmans ◽  
Andrew P. Halestrap ◽  
Urs T. Ruegg ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Reverse Transcription ◽  
Clinical Isolates ◽  
Ppiase Activity ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Ca Protein

ABSTRACT Debio-025 is a synthetic cyclosporine with no immunosuppressive capacity but a high inhibitory potency against cyclophilin A (CypA)-associated cis-trans prolyl isomerase (PPIase) activity. A lack of immunosuppressive effects compared to that of cyclosporine was demonstrated both in vitro and in vivo. For three cyclosporines, the inhibitory potential against PPIase activity was quantitatively correlated with that against human immunodeficiency virus type 1 (HIV-1) replication. Debio-025 selectively inhibited the replication of HIV-1 in a CD4+ cell line and in peripheral blood mononuclear cells: potent activity was demonstrated against clinical isolates of various HIV-1 subtypes, including isolates with multidrug resistance to reverse transcriptase and protease inhibitors. Simian immunodeficiency virus and HIV-2 strains were generally resistant to inhibition by Debio-025; however, some notable exceptions of sensitive HIV-2 clinical isolates were detected. In two-drug combination studies, additive inhibitory effects were found between Debio-025 and 19 clinically used drugs of different classes. Clinical HIV-1 isolates that are naturally resistant to Debio-025 and that do not depend on CypA for infection were identified. Comparison of the amino acid sequences of the CypA binding domain of the capsid (CA) protein from Debio-025-sensitive and -resistant HIV-1 isolates indicated that resistance was mostly associated with an H87Q/P exchange. Mechanistically, cyclosporines competitively inhibit the binding of CypA to the HIV-1 CA protein, which is an essential interaction required for early steps in HIV-1 replication. By real-time PCR we demonstrated that early reverse transcription is reduced in the presence of Debio-025 and that late reverse transcription is almost completely blocked. Thus, Debio-025 seems to interfere with the function of CypA during the progression/completion of HIV-1 reverse transcription.

scholarly journals Both Neutralization Resistance and High Infectivity Phenotypes Are Caused by Mutations of Interacting Residues in the Human Immunodeficiency Virus Type 1 gp41 Leucine Zipper and the gp120 Receptor- and Coreceptor-Binding Domains

1999 ◽  
Vol 73 (7) ◽  
pp. 5707-5713 ◽  
Author(s):  
Eun Ju Park ◽  
Gerald V. Quinnan
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Leucine Zipper ◽  
Vaccine Development ◽  
Viral Envelope ◽  
C Terminus ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Neutralization resistance of human immunodeficiency virus type 1 (HIV-1) is a major impediment to vaccine development. We have found that residues of HIV-1 MN strain in the C terminus of gp120 and the leucine zipper (LZ) region of gp41 viral envelope proteins interact cooperatively to determine neutralization resistance and modulate infectivity. Further, results demonstrate that this interaction, by which regions of gp120 are assembled onto the LZ, involves amino acid residues intimately related to those which participate in the binding of the envelope to its receptor and coreceptor. Variations in this critical assembly structure determine the concordant, interdependent evolution of increased infectivity efficiency and neutralization resistance phenotypes of the envelopes. The results elucidate important structure-function relationships among epitopes that are important targets of vaccine development.

scholarly journals Assembly Properties of the Human Immunodeficiency Virus Type 1 CA Protein

2004 ◽  
Vol 78 (5) ◽  
pp. 2545-2552 ◽  
Author(s):  
Barbie K. Ganser-Pornillos ◽  
Uta K. von Schwedler ◽  
Kirsten M. Stray ◽  
Christopher Aiken ◽  
Wesley I. Sundquist
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Hexagonal Lattice ◽  
Terminal Domain ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Ca Protein

ABSTRACT During retroviral maturation, the CA protein oligomerizes to form a closed capsid that surrounds the viral genome. We have previously identified a series of deleterious surface mutations within human immunodeficiency virus type 1 (HIV-1) CA that alter infectivity, replication, and assembly in vivo. For this study, 27 recombinant CA proteins harboring 34 different mutations were tested for the ability to assemble into helical cylinders in vitro. These cylinders are composed of CA hexamers and are structural models for the mature viral capsid. Mutations that diminished CA assembly clustered within helices 1 and 2 in the N-terminal domain of CA and within the crystallographically defined dimer interface in the CA C-terminal domain. These mutations demonstrate the importance of these regions for CA cylinder production and, by analogy, mature capsid assembly. One CA mutant (R18A) assembled into cylinders, cones, and spheres. We suggest that these capsid shapes occur because the R18A mutation alters the frequency at which pentamers are incorporated into the hexagonal lattice. The fact that a single CA protein can simultaneously form all three known retroviral capsid morphologies supports the idea that these structures are organized on similar lattices and differ only in the distribution of 12 pentamers that allow them to close. In further support of this model, we demonstrate that the considerable morphological variation seen for conical HIV-1 capsids can be recapitulated in idealized capsid models by altering the distribution of pentamers.

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