Structural Assembly of Qβ Virion and Its Diverse Forms of Virus-like Particles

Chang, Jeng-Yih; Gorzelnik, Karl V.; Thongchol, Jirapat; Zhang, Junjie

doi:10.3390/v14020225

Cited by 20 publications

(18 citation statements)

References 46 publications

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“…A model for MS2 assembly has been proposed in which upward of 60 strategically positioned packaging signals ( 18 , 47 ) guide the assembly of the coat proteins. While not providing direct evidence in support of this model, high-resolution cryoelectron microscopy studies of MS2 and Qβ particles show approximately one dozen specific interactions between the RNA genome and its surrounding protein capsid ( 31 , 32 , 48 – 50 ), suggesting that packaging signals play a role in ordering the genome. But BMV is different.…”

Section: Discussionmentioning

confidence: 98%

Single-particle studies of the effects of RNA–protein interactions on the self-assembly of RNA virus particles

Garmann

Goldfain

Tanimoto

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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Understanding the pathways by which simple RNA viruses self-assemble from their coat proteins and RNA is of practical and fundamental interest. Although RNA–protein interactions are thought to play a critical role in the assembly, our understanding of their effects is limited because the assembly process is difficult to observe directly. We address this problem by using interferometric scattering microscopy, a sensitive optical technique with high dynamic range, to follow the in vitro assembly kinetics of more than 500 individual particles of brome mosaic virus (BMV)—for which RNA–protein interactions can be controlled by varying the ionic strength of the buffer. We find that when RNA–protein interactions are weak, BMV assembles by a nucleation-and-growth pathway in which a small cluster of RNA-bound proteins must exceed a critical size before additional proteins can bind. As the strength of RNA–protein interactions increases, the nucleation time becomes shorter and more narrowly distributed, but the time to grow a capsid after nucleation is largely unaffected. These results suggest that the nucleation rate is controlled by RNA–protein interactions, while the growth process is driven less by RNA–protein interactions and more by protein–protein interactions and intraprotein forces. The nucleated pathway observed with the plant virus BMV is strikingly similar to that previously observed with bacteriophage MS2, a phylogenetically distinct virus with a different host kingdom. These results raise the possibility that nucleated assembly pathways might be common to other RNA viruses.

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Section: Discussionmentioning

confidence: 98%

Single-particle studies of the effects of RNA–protein interactions on the self-assembly of RNA virus particles

Garmann

Goldfain

Tanimoto

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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“…To test this hypothesis, future experiments and analysis might focus on the growth process rather than nucleation. Recently, BMV, MS2, and Qb capsids of different sizes and symmetries have been observed (40)(41)(42)(43)(44)(45). Experiments that determine how such malformed or overgrown capsids form would shed light on the interactions that control growth.…”

Section: Discussionmentioning

confidence: 99%

Single-particle studies of the effects of RNA-protein interactions on the self-assembly of RNA virus particles

Garmann

Goldfain²,

Tanimoto³

et al. 2022

Preprint

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Understanding the pathways by which simple RNA viruses self-assemble from their coat proteins and RNA is of practical and fundamental interest. Although RNA-protein interactions are thought to play a critical role in the assembly, our understanding of their effects is limited because the assembly process is difficult to observe directly. We address this problem by using interferometric scattering microscopy, a sensitive optical technique with high dynamic range, to follow the in vitro assembly kinetics of over 500 individual particles of brome mosaic virus (BMV)—for which RNA-protein interactions can be controlled by varying the ionic strength of the buffer. We find that when RNA-protein interactions are weak, BMV assembles by a nucleation-and-growth pathway in which a small cluster of RNA-bound proteins must exceed a critical size before additional proteins can bind. As the strength of RNA-protein interactions increases, the nucleation time becomes shorter and more narrowly distributed until the assembly kinetics become indistinguishable from diffusion-limited adsorption. In contrast, the time to grow a capsid after nucleation varies weakly with both salt and protein concentration. These results show that the nucleation rate is controlled by RNA-protein interactions, while the growth process is driven less by RNA-protein interactions and more by protein-protein interactions and intra-protein forces. The nucleated pathway observed with the plant virus BMV is strikingly similar to that previously observed with bacteriophage MS2, a phylogenetically distinct virus with a different host kingdom. These results raise the possibility that nucleated assembly pathways might be common to other RNA viruses.RNA viruses first inspired the term “self-assembly.” Yet much is still not understood about how even the simplest such viruses assemble or if different viruses assemble in similar ways. Theoretical models suggest many possible assembly pathways, with many different roles for the RNA, but until recently measuring these pathways has not been possible. We use a sensitive microscopy technique to follow the assembly of individual particles of BMV, a plant virus. We find evidence of an RNA-mediated nucleation-and-growth pathway that is strikingly similar to that of MS2, a bacterial virus. The last common ancestor of BMV and MS2 existed only in ancient times, suggesting that their assembly pathway might be evolutionarily conserved and other viruses might follow a similar pathway.

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“…There is no proline to potentially shift away interactions with surrounding residues, so this result suggests that hydrogen bonding is not the only feature that could be involved with forming the smaller sized phenotype in Qβ VLPs. Qβ CP[A40L/V41C] and Qβ CP[A40P/V41C] also both form Mini and WT VLPs, but both variants also seem to form a prolate shaped VLP which forms as an elongated particle VLPs (Figure 8 C-D) [12].…”

Section: Generating and Assessing A 2d-afl For Qβ 40/41mentioning

confidence: 99%

“…Qβ bacteriophages form small percentages of mini-sized assembled capsids naturally even if those variants are not infective and therefore are less than 5% of the particle population [12]. Crystal structure analysis of the VLP has shown that Qβ dimer conformations are flexible, being that the crucial VLP stabilizing interactions are intradimer salt bridges and other amino acid interactions.…”

Section: Generating and Assessing A 2d-afl For Qβ 40/41mentioning

confidence: 99%

“…For example, a double variant of the tobacco mosaic virus (TMV) coat protein (CP) yielded a “nanodisk”[11] assembly instead of the standard rod assembly. Single variants in the CP of the bacteriophage Qβ yielded a size distribution of assemblies that included smaller, wild-type (WT)-sized, and longer prolate VLPs [12, 13]. A single amino acid variant in bacteriophage MS2 CP [S37P] (“Mini MS2”) [14] maintains nearly identical protein sequence, and has remarkably similar secondary and tertiary structure to the parent CP; however, the quaternary structure changes dramatically.…”

Section: Introductionmentioning

confidence: 99%

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Systematic Engineering of Virus-Like Particles to Identify Self-Assembly Rules for Shifting Particle Size

Ikwuagwu

Hartman

Mills

et al. 2022

Preprint

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Virus-like particles (VLPs) are promising scaffolds for biomaterials as well as diagnostic and therapeutic applications. However, there are some key challenges to be solved, such as the ability to engineer alternate sizes for varied use cases. To this end, we created a library of MS2 VLP variants at two key residues in the coat protein which have been implicated as important to controlling VLP size and geometry. By adapting a method for systematic mutagenesis coupled with size-based selections and high-throughput sequencing as a readout, we developed a quantitative assessment of two residues in MS2 coat protein that govern the size shift in MS2 VLPs. We then applied the strategy to the equivalent residues in Qβ VLPs, an MS2 homolog, and demonstrate that the analogous pair of residues are also able to impact VLP size and shape. These results underscore the power of fitness landscapes in identifying critical features for assembly.

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Structural Assembly of Qβ Virion and Its Diverse Forms of Virus-like Particles

Cited by 20 publications

References 46 publications

Single-particle studies of the effects of RNA–protein interactions on the self-assembly of RNA virus particles

Single-particle studies of the effects of RNA–protein interactions on the self-assembly of RNA virus particles

Single-particle studies of the effects of RNA-protein interactions on the self-assembly of RNA virus particles

Systematic Engineering of Virus-Like Particles to Identify Self-Assembly Rules for Shifting Particle Size

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