Towards the structure of the human immunodeficiency virus: divide and conquer?

Self Cite

164

162

Immature retrovirus particles contain radially arranged Gag polyproteins in which the N termini lie at the membrane and the C termini extend toward the particle's center. We related image features to the polyprotein domain structure by combining mutagenesis with cryoelectron microscopy and image analysis. The matrix (MA) domain appears as a thin layer tightly associated with the inner face of the viral membrane, separated from the capsid (CA) layer by a low-density region corresponding to its C terminus. Deletion of the entire p6 domain has no effect on the width or spacing of the density layers, suggesting that p6 is not ordered in immature human immunodeficiency virus type 1 (HIV-1). In vitro assembly of a recombinant Gag polyprotein containing only capsid (CA) and nucleocapsid (NC) domains results in the formation of nonenveloped spherical particles which display two layers with density matching that of the CA-NC portion of immature HIV-1 Gag particles. Authentic, immature HIV-1 displays additional surface features and an increased density between the lipid bilayers which reflect the presence of gp41. The other internal features match those of virus-like particles.

Section: Resultsmentioning

confidence: 82%

Section: The Genome Of the Human Immunodeficiency Virus (Hiv)mentioning

confidence: 98%

Section: The Genome Of the Human Immunodeficiency Virus (Hiv)mentioning

confidence: 99%

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Organization of Immature Human Immunodeficiency Virus Type 1

Wilk

Gross

Gowen

et al. 2001

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164

162

“…FV share the heterogeneous size of released retroviral particles, the lack of icosahedral symmetry, and the radial arrangement of defined Gag domains with the other retroviruses that have been examined (20,47,49,52).…”

Section: Discussionmentioning

confidence: 99%

Specific Interaction of a Novel Foamy Virus Env Leader Protein with the N-Terminal Gag Domain

Wilk

Geiselhart

Frech

et al. 2001

Self Cite

125

Cryoelectron micrographs of purified human foamy virus (HFV) and feline foamy virus (FFV) particles revealed distinct radial arrangements of Gag proteins. The capsids were surrounded by an internal Gag layer that in turn was surrounded by, and separated from, the viral membrane. The width of this layer was about 8 nm for HFV and 3.8 nm for FFV. This difference in width is assumed to reflect the different sizes of the HFV and FFV MA domains: the HFV MA domain is about 130 residues longer than that of FFV. The distances between the MA layer and the edge of the capsid were identical in different particle classes. In contrast, only particles with a distended envelope displayed an invariant, close spacing between the MA layer and the Env membrane which was absent in the majority of particles. This indicates a specific interaction between MA and Env at an unknown step of morphogenesis. This observation was supported by surface plasmon resonance studies. The purified N-terminal domain of FFV Gag specifically interacted with synthetic peptides and a defined protein domain derived from the N-terminal Env leader protein. The specificity of this interaction was demonstrated by using peptides varying in the conserved Trp residues that are known to be required for HFV budding. The interaction with Gag required residues within the novel virion-associated FFV Env leader protein of about 16.5 kDa. The genomes of spumaretroviruses (or foamy viruses [FV])include the classical gag, pro-pol, and env genes that are the hallmark of the retrovirus family (9). Despite the familial relationship implied by their genome organization, FV differ from retroviruses such as oncoviruses or lentiviruses in basic aspects of replication and gene expression (30-32, 39, 53). The differences between FV and the more widely studied retroviruses, such as human immunodeficiency virus (HIV) or murine leukemia virus, provide an opportunity to identify the fundamental mechanisms of processes such as particle assembly and maturation.The shared gene order of the FV genomes and those of other retroviruses allows the identification of common structural proteins. Unfortunately, this relationship does not extend to the structure and function of FV Gag proteins, since no obvious homologies are detectable between FV Gag and MA, CA, and NC domains of other retroviruses for which structures have been determined to high resolution (10, 43). Furthermore, the proteolytic processing of Gag that produces the familiar mature proteins in other retroviruses is unusual, incomplete, and/or delayed in FV (12,23,29,37).The available data indicate that the morphology and morphogenesis of FV are distinct from those of other retroviruses. FV Gag proteins preassemble to form in the cytoplasm spherical capsids that bud through cellular membranes (55). This process contrasts with what occurs with lentiviruses and C-type retroviruses that assemble their capsids at the site of budding but is similar to what occurs with B-and D-type retroviruses (40). Budding of human foamy virus (HFV) i...

“…In addition, an N-terminal mutation of murine leukemia virus CA functions in a transdominant manner (33). Most HIV-1 proteins function in a multimeric complex, although the virion complex of several thousand Gag proteins is by far the largest complex among the viral proteins (7,46). This suggests that mutant Gag proteins should have the potential to display strong multiplicative effects on their inhi-bition of virion infectivity.…”

mentioning

confidence: 99%

A Strongly Transdominant Mutation in the Human Immunodeficiency Virus Type 1 gag Gene Defines an Achilles Heel in the Virus Life Cycle

Lee

Harris

Swanstrom

2009

114

The human immunodeficiency virus type 1 (HIV-1) protease (PR) makes five obligatory cleavages in the viral Gag polyprotein precursor. The cleavage events release the virion structural proteins from the precursor and allow the virion to undergo maturation to become infectious. The protease cleavage between the matrix protein (MA) domain and the adjacent capsid protein (CA) domain releases CA from the membrane-anchored MA and allows the N terminus of CA to refold into a structure that facilitates the formation of hexamer arrays that represent the structural unit of the capsid shell. In this study, we analyzed the extent to which each of the HIV-1 Gag processing sites must be cleaved by substituting the P1-position amino acid at each processing site with Ile. A mutation that blocks cleavage at the MA/CA processing site (Y132I) displayed a strong transdominant effect when tested in a phenotypic mixing strategy, inhibiting virion infectivity with a 50% inhibitory concentration of only 4% of the mutant relative to the wild type. This mutation is 10-to 20-fold more potent in phenotypic mixing than an inactivating mutation in the viral protease, the target of many successful inhibitors, and more potent than an inactivating mutation at any of the other Gag cleavage sites. The transdominant effect is manifested as the assembly of an aberrant virion core. Virus containing 20% of the Y132I mutant and 80% of the wild type (to assess the transdominant effect on infectivity) was blocked either before reverse transcription (RT) or at an early RT step. The ability of a small amount of the MA/CA fusion protein to poison the oligomeric assembly of infectious virus identifies an essential step in the complex process of virion formation and maturation. The effect of a small-molecule inhibitor that is able to block MA/CA cleavage even partially would be amplified by this transdominant negative effect on the highly orchestrated process of virion assembly.