In natural infections with hepatitis B virus (HBV), there is an excess of empty noninfectious subviral particles (SVP) that do not contain the viral capsid. SVP are typically present in a 1,000- to 100,000-fold excess relative to the infectious particles (12
). They exist in two main forms: spheres of 25 nm in diameter and filaments of 22 nm in diameter with variable length (15
). They can contain all three forms of the HBV envelope proteins: L, M, and S. These share a common C terminus, with M containing the pre-S2 domain relative to S and L containing the pre-S1 domain relative to M (15
). There is good evidence that during infection a domain within the pre-S1 of L is what interacts with an as-yet-unidentified host receptor(s) (15
). Hepatitis delta virus (HDV), which is assembled using the envelope proteins of HBV, also depends upon this pre-S1 domain (26
). HDV can be assembled using only the S protein of HBV, but the particles are noninfectious.
SVP from patients are immunogenic and were used with success in the first HBV vaccine (1
). Most current HBV vaccines are prepared in yeast from SVP assembled using just the HBV S protein; such SVP are sufficient to protect individuals against HBV and HDV.
The basis for the excess of SVP detected in patients is unexplained, and the biological function of this excess has been largely ignored. Some authors have suggested that SVP might sop up neutralizing antibodies produced by the host and thus increase the ability of the infectious particles to reach susceptible cells (11
). It has also been suggested that SVP contribute to a state of immune tolerance that is a precondition for highly productive persistent infection (13
). One study with SVP of duck HBV indicated that for infections at low multiplicity SVP could enhance infection, but when present in large amounts they were inhibitory (3
). Another study showed that SVP containing the large envelope protein interfered with duck HBV infection (19
For the present studies we chose to use SVP as produced by transfection procedures. For the following reasons we consider these more defined than SVP obtained from the sera of infected individuals: (i) the latter contain infectious virus as well as SVP; (ii) they may also contain a spectrum of host antibodies either mixed with the SVP or even directly attached to the SVP; and (iii) since the patients producing SVP are chronically infected, the genetic composition of the SVP will be mixed, with a variety of mutant forms. In contrast, with the in vitro approach, we can assemble SVP that contain just the HBV S protein or those with both the S and L proteins.
As described below, we assembled not just different forms of SVP but also HBV and HDV. We thus found that during HBV and HDV assembly, there was not necessarily a great excess of SVP. In addition, we assembled SVP in the absence of HBV and HDV and found that these SVP were able to bind heparin in vitro yet were not able to interfere with the infection of primary human hepatocytes (PHH) by HBV or HDV.