Native display of complete foreign protein domains on the surface of hepatitis B virus capsids

The nucleocapsid of hepatitis B virus (HBV), or HBcAg, is a highly symmetri c structure formed by multiple dimers of a single core protein that contain s potent T helper epitopes in its 183-aa sequence. Both factors make HBcAg an unusually strong immunogen and an attractive candidate as a carrier for foreign epitopes. The immunodominant c/e1 epitope on the capsid has been su ggested as a superior location to convey high immunogenicity to a heterolog ous sequence. Because of its central position, however, any c/e1 insert dis rupts the core protein's primary sequence; hence, only peptides, or rather small protein fragments seemed to be compatible with particle formation. Ac cording to recent structural data, the epitope is located at the tips of pr ominent surface spikes formed by the very stable dimer interfaces. We there fore reasoned that much larger inserts might be tolerated, provided the ind ividual parts of a corresponding fusion protein could fold independently. U sing the green fluorescent protein (GFP) as a model insert, we show that th e chimeric protein efficiently forms fluorescent particles; hence, all of i ts structurally important parts must be properly folded. We also demonstrat e that the GFP domains are surface-exposed and that the chimeric particles elicit a potent humoral response against native GFP. Hence, proteins of at least up to 238 aa can be natively displayed on the surface of HBV core par ticles. Such chimeras may not only be useful as vaccines but may also open the way for high resolution structural analyses of non-assembling proteins by electron microscopy.