Structure and Stoichiometry of Template-Directed Recombinant HIV-1 Gag Particles

Goicochea, Nancy L.; Datta, Siddhartha A.K.; Murali, Ayaluru; Kao, C. Cheng; Rein, Alan; Dragnea, Bogdan

doi:10.1016/j.jmb.2011.04.012

Cited by 22 publications

(30 citation statements)

References 54 publications

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“…[36] The use of a template significantly reduced the size polydispersity of the assembled VLPs. Specifically, 60 nm gold nanoparticles were found to be the most efficient for generating homogeneous Gag VLPs.…”

Section: Introductionmentioning

confidence: 99%

Virus Matryoshka: A Bacteriophage Particle—Guided Molecular Assembly Approach to a Monodisperse Model of the Immature Human Immunodeficiency Virus

Saxena

Malyutin

et al. 2016

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Immature human immunodeficiency virus type 1 (HIV-1) is approximately spherical, but is constructed from a hexagonal lattice of the Gag protein. As a hexagonal lattice is necessarily flat, the local symmetry cannot be maintained throughout the structure. This geometrical frustration presumably results in bending stress. In natural particles, the stress is relieved by incorporation of packing defects, but the magnitude of this stress and its significance for the particles is not known. In order to control this stress, we have now assembled the Gag protein on a quasi-spherical template derived from bacteriophage P22. This template is monodisperse in size and electron-transparent, enabling the use of cryo-electron microscopy in structural studies. These templated assemblies are far less polydisperse than any previously described virus-like particles (and, while constructed according to the same lattice as natural particles, contain almost no packing defects). This system gives us the ability to study the relationship between packing defects, curvature and elastic energy, and thermodynamic stability. As Gag is bound to the P22 template by single-stranded DNA, treatment of the particles with DNase enabled us to determine the intrinsic radius of curvature of a Gag lattice, unconstrained by DNA or a template. We found that this intrinsic radius is far larger than that of a virion or P22-templated particle. We conclude that Gag is under elastic strain in a particle; this has important implications for the kinetics of shell growth, the stability of the shell, and the type of defects it will assume as it grows.

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Section: Introductionmentioning

confidence: 99%

Virus Matryoshka: A Bacteriophage Particle—Guided Molecular Assembly Approach to a Monodisperse Model of the Immature Human Immunodeficiency Virus

Saxena

Malyutin

et al. 2016

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“…49 The HIV-1 Gag-VLPs were chosen to test the predictions from numerical simulations because they are believed to exhibit sizeable gaps in their protein lattice. 49 The existence of gaps in the HIV-1 immature particles and HIV-1 Gag-VLPs were previously determined by electron tomography, 57, 64 but never by any other method. The current view on the native Gag shell is that of a lattice with openings shaped as scars resulting in a total surface coverage of ~70%.…”

Section: Resultsmentioning

confidence: 99%

“…49 The ~25% signal increase from the insulation due to the protein layer corresponds then to a contribution of ~ 0.008% per protein and 0.05% per hexameric subunit. This is below the noise level of the current instrument, at least for temperatures that are reasonable for in vivo conditions.…”

Section: Resultsmentioning

confidence: 99%

“… a COMSOL parameter databank, b calculated according to D=κ/ρC p c calculated from molar weight of the ligands, #of ligands per Au NP estimated from Hurst et al 48 and volume in TEM and DLS experiments, d calculated from molar weight of the proteins, average # of proteins 49 , e sum of contributions from DNA 50 and tetraethylene glycol 51 , f sum of contributions of the molar heat capacities of the comprising amino acids 52 , g estimated from mass mixture of the ligands 53, 54, 55 , h estimated from protein volume 56 …”

Section: Figurementioning

confidence: 99%

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Photothermal Imaging and Measurement of Protein Shell Stoichiometry of Single HIV-1 Gag Virus-like Nanoparticles

et al. 2011

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Virus life stages often constitute a complex chain of events, difficult to track in-vivo and in real-time. Challenges are associated with spatial and time limitations of current probes: most viruses are smaller than the diffraction limit of optical microscopes while the entire time-scale of virus dynamics spans over 8 orders of magnitude. Thus, virus processes such as entry, disassembly, and egress have generally remained poorly understood. Here we discuss photothermal heterodyne imaging (PHI) as a possible alternative to fluorescence microscopy in the study of single virus-like nanoparticle (VNP) dynamics, with relevance in particular to virus uncoating. Being based on optical absorption rather than emission, PHI could potentially surpass some of the current limitations associated with fluorescent labels. As proof-of-principle, single VNPs self-assembled from 60 nm DNA-functionalized gold nanoparticles (DNA-Au NPs) encapsulated in a Gag protein shell of the Human immunodeficiency virus (HIV-1) were imaged and their photothermal response compared with DNA-Au NPs. For the first-time, the protein stoichiometry of a single virus-like particle was estimated by a method other than electron microscopy.

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“…In this study, we have examined the feasibility of assembling proteinaceous AaLS‐13 shells around inorganic nanomaterials . Spherical gold nanoparticles (AuNPs) are useful for drug delivery, for photocatalysis, as biological sensors, and as conjugation platforms for oligonucleotides and biomolecules .…”

Section: Figurementioning

confidence: 99%

Self‐Assembly of Proteinaceous Shells around Positively Charged Gold Nanomaterials Enhances Colloidal Stability in High‐Ionic‐Strength Buffers

et al. 2019

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The enzyme lumazine synthase (LS) has been engineered to self‐assemble into hollow‐shell structures that encapsulate unnatural cargo proteins through complementary electrostatic interactions. Herein, we show that a negatively supercharged LS variant can also form organic–inorganic hybrids with gold nanomaterials. Simple mixing of LS pentamers with positively charged gold nanocrystals in aqueous buffer spontaneously affords protein‐shelled gold cores. The procedure works well with differently sized and shaped gold nanocrystals, and the resulting shelled complexes exhibit dramatically enhanced colloidal stability over a wide range of pH (4.0–10.0) and at high ionic strength (up to 1 m NaCl). They are even stable over days upon dilution in buffer. Self‐assembly of engineered LS shells in this way offers an easy and attractive alternative to commonly used ligand‐exchange methods for stabilizing inorganic nanomaterials.

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Structure and Stoichiometry of Template-Directed Recombinant HIV-1 Gag Particles

Cited by 22 publications

References 54 publications

Virus Matryoshka: A Bacteriophage Particle—Guided Molecular Assembly Approach to a Monodisperse Model of the Immature Human Immunodeficiency Virus

Virus Matryoshka: A Bacteriophage Particle—Guided Molecular Assembly Approach to a Monodisperse Model of the Immature Human Immunodeficiency Virus

Photothermal Imaging and Measurement of Protein Shell Stoichiometry of Single HIV-1 Gag Virus-like Nanoparticles

Self‐Assembly of Proteinaceous Shells around Positively Charged Gold Nanomaterials Enhances Colloidal Stability in High‐Ionic‐Strength Buffers

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