Rapidly Forming Early Intermediate Structures Dictate the Pathway of Capsid Assembly

Roi Asor, Christopher John Schlicksup, Zhongchao Zhao, Adam Zlotnick, Uri Raviv*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

29 Scopus citations


There are 1030 possible intermediates on the assembly path from hepatitis B capsid protein dimers to the 120-dimer capsid. If every intermediate was tested, assembly would often get stuck in an entropic trap and essentially every capsid would follow a unique assembly path. Yet, capsids assemble rapidly with minimal trapped intermediates, a realization of the Levinthal paradox. To understand the fundamental mechanisms of capsid assembly, it is critical to resolve the early stages of the reaction. We have used time-resolved small angle X-ray scattering, which is sensitive to solute size and shape and has millisecond temporal resolution. Scattering curves were fit to a thermodynamically curated library of assembly intermediates, using the principle of maximum entropy. Maximum entropy also provides a physical rationale for the selection of species. We found that the capsid assembly pathway was exquisitely sensitive to initial assembly conditions. With the mildest conditions tested, the reaction appeared to be two-state from dimer to 120-dimer capsid with some dimers-of-dimers and trimers-of-dimers. In slightly more aggressive conditions, we observed transient accumulation of a decamer-of-dimers and the appearance of 90-dimer capsids. In conditions where there is measurable kinetic trapping, we found that highly diverse early intermediates accumulated within a fraction of a second and propagated into long-lived kinetically trapped states (≥90-mer). In all cases, intermediates between 35 and 90 subunits did not accumulate. These results are consistent with the presence of low barrier paths that connect early and late intermediates and direct the ultimate assembly path to late intermediates where assembly can be paused.

Original languageAmerican English
Pages (from-to)7868-7882
Number of pages15
JournalJournal of the American Chemical Society
Issue number17
StatePublished - 29 Apr 2020

Bibliographical note

Funding Information:
We thank Daniel Harries for very fruitful and enlightening discussions and Corinne A. Lutomski and Martin F. Jarrold for sharing their CDMS data, presented in Figure S6. We acknowledge the European Synchrotron Radiation Facility (ESRF) beamline ID02 (T. Narayanan and his team), the Desy synchrotron at Hamburg, beamline P12 (D. Svergun and his team), and Soleil synchrotron, Swing beamline (J. Perez and his team), for provision of synchrotron radiation facilities and for assistance in using the beamlines. This project was supported by the NIH (Award Number 1R01AI118933 to A.Z.). R.A. acknowledges support from the Kaye-Einstein Fellowship Foundation. U.R. and R.A. acknowledge financial support from the Israel Science Foundation (Grant 656/17).

Publisher Copyright:
Copyright © 2020 American Chemical Society.


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