Hepatitis virus capsid polymorph stability depends on encapsulated cargo size

Abstract

Protein cages providing a controlled environment to encapsulated cargo are a ubiquitous presence in any biological system. Well-known examples are capsids, the regular protein shells of viruses, which protect and deliver the viral genome. Since some virus capsids can be loaded with nongenomic cargoes, they are interesting for a variety of applications ranging from biomedical delivery to energy harvesting. A question of vital importance for such applications is how does capsid stability depend on the size of the cargo? A nanoparticle-templated assembly approach was employed here to determine how different polymorphs of the Hepatitis B virus icosahedral capsid respond to a gradual change in the encapsulated cargo size. It was found that assembly into complete virus-like particles occurs cooperatively around a variety of core diameters, albeit the degree of cooperativity varies. Among these virus-like particles, it was found that those of an outer diameter corresponding to an icosahedral array of 240 proteins (T = 4) are able to accommodate the widest range of cargo sizes.

Figure 1

Schematic principle of testing the possibility for differential response of capsid polymorphs to an increase in the cargo size. a) closed capsid resisting to an increase in cargo diameter. b) CP shell adapting to increase in cargo size by an increase in the outer diameter facilitated by packing defects (opening).

Figure 2

a) Completely formed VLPs and bare Au NPs are the two main states in which a Au NP can be found at CP concentrations below NP surface saturation value. Scale bar: 100 nm. b) TEM picture of VLP encapsulating a 15 nm NP core is suggestive of a regular array of CPs. c) Pie-chart showing relative frequencies of complete VLPs (black), incomplete VLPs (dark gray), and bare NPs for an initial CP:NP molar ratio of 50.

Figure 3

a) Assembly efficiency as a function of CP:NP molar ratio for the two core sizes which correspond to the T=3 and T=4 HBV capsid lumen diameters, respectively (error bars from replicates). b) Same data but shown against estimated free CP molar concentration, x. Lines represent Hill plots (eq. 1). c) A Langmuir model (dotted line) cannot fit the 19 nm data.

Figure 4

Fractions of complete and incomplete VLPs from a competition experiment in conditions of limited CP supply. Red dots: expected from non-cooperative adsorption.

Figure 5

Efficiency of encapsulation of 15 nm diameter NPs as a function of PEG-COOH coverage and number of positive charges per CP tail.

Figure 6

Probability density map of VLP diameters resulting from certain NP core diameters. The dotted line represents the increase in VLP diameter with NP core diameter in conditions of a hypotethical constant protein layer thickness of 7.5 nm.