Abstract
Self-assembly is a vital part of the life cycle of certain icosahedral RNA viruses. Furthermore, the assembly process can be harnessed to make icosahedral virus-like particles (VLPs) from coat protein and RNA in vitro. Although much previous work has explored the effects of RNA-protein interactions on the assembly products, relatively little research has explored the effects of coat-protein concentration. We mix coat protein and RNA from bacteriophage MS2, and we use a combination of gel electrophoresis, dynamic light scattering, and transmission electron microscopy to investigate the assembly products. We show that with increasing coat-protein concentration, the products transition from well-formed MS2 VLPs to "monster" particles consisting of multiple partial capsids to RNA-protein condensates consisting of large networks of RNA and partially assembled capsids. We argue that the transition from well-formed to monster particles arises because the assembly follows a nucleation-and-growth pathway in which the nucleation rate depends sensitively on the coat-protein concentration, such that at high protein concentrations, multiple nuclei can form on each RNA strand. To understand the formation of the condensates, which occurs at even higher coat-protein concentrations, we use Monte Carlo simulations with coarse-grained models of capsomers and RNA. These simulations suggest that the formation of condensates occurs by the adsorption of protein to the RNA followed by the assembly of capsids. Multiple RNA molecules can become trapped when a capsid grows from capsomers attached to two different RNA molecules or when excess protein bridges together growing capsids on different RNA molecules. Our results provide insight into an important biophysical process and could inform design rules for making VLPs for various applications.
| Original language | English |
|---|---|
| Pages (from-to) | 3121-3132 |
| Number of pages | 12 |
| Journal | Nanoscale |
| Volume | 16 |
| Issue number | 6 |
| Early online date | 17 Jan 2024 |
| DOIs | |
| Publication status | Published - 14 Feb 2024 |
Bibliographical note
Acknowledgments:We thank Amy Barker and Peter Stockley at the University of Leeds for initial stocks of MS2 and E. coli cells. We thank Tim Chiang, Amelia Paine, Aaron Goldfain, and Danai Montalvan for helpful scientific discussions. This research was partially supported by a National Science Foundation (NSF) Graduate Research Fellowship under grant number DGE-1745303, by NSF through the Harvard University Materials Research Science and Engineering Center under NSF grant number DMR-2011754, by the National Institute of General Medical Sciences of the National Institutes of Health under grant numbers K99GM127751 and R00GM127751, by the NSF-Simons Center for Mathematical and Statistical Analysis of Biology at Harvard University under NSF grant number 1764269, and by the Harvard Quantitative Biology Initiative. AN, VNM, and DC gratefully acknowledge support from the Institute of Advanced Studies of the University of Birmingham and the Turing Scheme. This work was performed in part at the Harvard University Center for Nanoscale Systems (CNS), a member of the National Nanotechnology Coordinated Infrastructure Network (NNCI), which is supported by the National Science Foundation under NSF grant number ECCS-2025158. The work was also performed in part at the Harvard University Bauer Core Facility. Any opinion, findings, and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of the National Science Foundation.
Keywords
- Capsid
- Levivirus/genetics
- Capsid Proteins/metabolism
- RNA, Viral/genetics
- Virion