Interaction of Hepatitis B capsid like particles with surface protein fragments and peptides interfering with envelopment
Final Report Abstract
Hepatitis B Virus is a major human pathogen with some 250 Mio carriers worldwide. The virions have an icosahedral capsid with protruding spikes. The capsids are surrounded by a pleiomorphic lipidic envelope that is densely packed with surface proteins The envelopment of the capsids is crucial for secretion of mature virions and for infectivity. Disrupting the interaction between surface proteins and capsid proteins is therefore a potential route to an antiviral drug. The interaction site between capsids and envelope is mapped to the spikes, but it is still controversial whether the surface proteins bind to the tips of the spikes or to a pocket in the centre of the spikes. We investigated the binding of surface protein fragments to capsids and found that they bind very weakly and do not decorate the capsids consistently. We obtained evidence that avidity of multiple interlinked binding sites is required to observe an interaction between the capsids and the surface proteins. In contrast peptides that interfere with the envelopment bind much more strongly and decorate the tips of the spikes consistently. Their binding splays the protruding part of the spikes apart without stabilizing the spike structure. This did not change in capsid mutants with either a low (P5T, L60V) or a premature (F97L) secretion phenotype. The other potential binding site for surface proteins is in the middle of the spikes. We found that this site binds small hydrophobic molecules, some of them are mimetics for either prenylation or for myristylation. The strongest of the tested binders had an aromatic moiety and altered the rotamer conformation of residue F97 in the capsid protein. The effect was similar for the other mutants (F97L, P5T and L60V). Both binding sites at the spike could be addressed simultaneously. The binding modes of the peptides and small molecules did not correlate with the reported secretion phenotypes of the capsids. Based on our binding studies, we designed super-binders with increased affinity. The best binder had nanomolar affinity and reduced the capsid solubility. Taking advantage of this property, we developed a binder that entered cells and aggregated capsids inside these cells. We believe that this binder can be further developed into a tool for targeting and removing capsids from the viral maturation cycle at an early stage by phase separation.
Publications
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Protein Data Bank (PDB): 7OCO (L60V-HBc), 7OCW (P5T-HBc), 7OD4 (WT-HBc), 7OD6 (WT-HBc + P2), 7OD7 (WT+SLLGRM), 7OD8 (L60V-HBc + P2)
Makbul et al.
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Protein Data Bank (PDB): PDB-7PZ9 (HBc-F97L), PDB-7PZI (HBc-F97L+TX100), PDB- 7PZK (HBc +TX100), PDB-7PZN (HBc +TX100+SLLGRM), PDB-7PZL (HBc-L60V +TX100), PDB-7PZM (HBc-P5T +TX100)
Makbul et al.
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Binding of a Pocket Factor to Hepatitis B Virus Capsids Changes the Rotamer Conformation of Phenylalanine 97. Viruses, 13(11), 2115.
Makbul, Cihan; Kraft, Christian; Grießmann, Matthias; Rasmussen, Tim; Katzenberger, Kilian; Lappe, Melina; Pfarr, Paul; Stoffer, Cato; Stöhr, Mara; Wandinger, Anna-Maria & Böttcher, Bettina
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Conformational Plasticity of Hepatitis B Core Protein Spikes Promotes Peptide Binding Independent of the Secretion Phenotype. Microorganisms, 9(5), 956.
Makbul, Cihan; Khayenko, Vladimir; Maric, Hans Michael & Böttcher, Bettina
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Electron Microscopy Data Bank: EMD-12810 (L60V-HBc), EMD-12815 (P5T-HBc), EMD-12819 (WT-HBc), EMD-12820 (WT-HBc+P2), EMD-12821(WT-HBc + ‘SLLGRM’), EMD-12822 (L60V-HBc + P2), EMD-12823 (F97L-HBc+P1), EMD-12824 (F97L- HBc + P2), EMD-12825, (P5T-HBc + P2).
Makbul et al.
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EMD-13726 (HBc-F97L), EMD-13728 (HBc-F97L+TX100), EMD-13731 (HBc +TX100), EMD-13734 (HBc +TX100+SLLGRM), EMD-13732 (HBc-L60V +TX100), EMD-13733 (HBc-P5T +TX100).
Makbul et al.
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Induction of Hepatitis B Core Protein Aggregation Targeting an Unconventional Binding Site. eLife Sciences Publications, Ltd.
Khayenko, Vladimir; Makbul, Cihan; Schulte, Clemens; Hemmelmann, Naomi; Kachler, Sonja; Böttcher, Bettina & Maric, Hans M.
