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An N-terminal extension to the hepatitis B virus core protein forms a poorly ordered trimeric spike in assembled virus-like particles.

McGonigle R, Yap WB, Ong ST, Gatherer D, Bakker SE, Tan WS, Bhella D - J. Struct. Biol. (2014)

Bottom Line: Heterologous sequences have been successfully inserted at both amino and carboxy termini as well as internally at the major immunodominant epitope.The insert was seen to form a trimeric spike on the capsid surface that was poorly resolved, most likely owing to it being flexible.We hypothesise that the capacity of N-terminal inserts to form trimers may have application in the development of multivalent vaccines to trimeric antigens.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Sir Michael Stoker Building, Garscube Campus, 464 Bearsden Road, Glasgow G61 1QH, Scotland, UK.

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3D reconstructions of His-β-L HBcAg virus-like particles. (A) Cryomicroscopy of His-β-L HBcAg VLPs revealed two sizes 30 nm T = 3 and 34 nm T = 4. (B) The T = 3 structure was generated using 6048 particle images and achieved a resolution of 10 Å. (C) The T = 4 VLP was reconstructed using 2040 particles and achieved a resolution of 12 Å. Resolution assessment was by the FSC0.5 criterion (D). The general architecture of both capsids is similar to the wild type HBcAg, as demonstrated by comparison with a solvent exclusion surface representation of the T = 4 capsid calculated from the X-ray structure PDB ID 1QGT (E). His-β-L HBcAg particles however were seen to bear an extra spike density hypothesised to comprise a trimer of the extended N-terminus, located at sites of local threefold symmetry in the T = 3 VLP (magenta) and at both icosahedral (green) and local (magenta) threefold symmetry axes in the T = 4 particle. Wall-eyed stereo pair images are presented (B, C and E). Cross-sections through the T = 3 (F) and T = 4 (G) reconstructions show that the density in the N-terminal spikes is blurred compared to that of the HBcAg component (magenta and green arrows).
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f0005: 3D reconstructions of His-β-L HBcAg virus-like particles. (A) Cryomicroscopy of His-β-L HBcAg VLPs revealed two sizes 30 nm T = 3 and 34 nm T = 4. (B) The T = 3 structure was generated using 6048 particle images and achieved a resolution of 10 Å. (C) The T = 4 VLP was reconstructed using 2040 particles and achieved a resolution of 12 Å. Resolution assessment was by the FSC0.5 criterion (D). The general architecture of both capsids is similar to the wild type HBcAg, as demonstrated by comparison with a solvent exclusion surface representation of the T = 4 capsid calculated from the X-ray structure PDB ID 1QGT (E). His-β-L HBcAg particles however were seen to bear an extra spike density hypothesised to comprise a trimer of the extended N-terminus, located at sites of local threefold symmetry in the T = 3 VLP (magenta) and at both icosahedral (green) and local (magenta) threefold symmetry axes in the T = 4 particle. Wall-eyed stereo pair images are presented (B, C and E). Cross-sections through the T = 3 (F) and T = 4 (G) reconstructions show that the density in the N-terminal spikes is blurred compared to that of the HBcAg component (magenta and green arrows).

Mentions: Cryomicrographs of purified His-β-L HBcAg VLPs showed the presence of two populations of differently sized particles with diameters of approximately 30 nm and 34 nm, corresponding to the expected sizes for T = 3 and T = 4 classes (Fig. 1A). The T = 3 form was most common (74.4%), in line with the findings of previous studies showing that truncation of the C-terminus increases the ratio of T = 3 versus T = 4 particles (Pumpens and Grens, 2001). 7046 T = 3 and 2419 T = 4 particles were extracted from 785 micrographs and processed to calculate three-dimensional reconstructions.


An N-terminal extension to the hepatitis B virus core protein forms a poorly ordered trimeric spike in assembled virus-like particles.

McGonigle R, Yap WB, Ong ST, Gatherer D, Bakker SE, Tan WS, Bhella D - J. Struct. Biol. (2014)

3D reconstructions of His-β-L HBcAg virus-like particles. (A) Cryomicroscopy of His-β-L HBcAg VLPs revealed two sizes 30 nm T = 3 and 34 nm T = 4. (B) The T = 3 structure was generated using 6048 particle images and achieved a resolution of 10 Å. (C) The T = 4 VLP was reconstructed using 2040 particles and achieved a resolution of 12 Å. Resolution assessment was by the FSC0.5 criterion (D). The general architecture of both capsids is similar to the wild type HBcAg, as demonstrated by comparison with a solvent exclusion surface representation of the T = 4 capsid calculated from the X-ray structure PDB ID 1QGT (E). His-β-L HBcAg particles however were seen to bear an extra spike density hypothesised to comprise a trimer of the extended N-terminus, located at sites of local threefold symmetry in the T = 3 VLP (magenta) and at both icosahedral (green) and local (magenta) threefold symmetry axes in the T = 4 particle. Wall-eyed stereo pair images are presented (B, C and E). Cross-sections through the T = 3 (F) and T = 4 (G) reconstructions show that the density in the N-terminal spikes is blurred compared to that of the HBcAg component (magenta and green arrows).
© Copyright Policy - CC BY
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4318616&req=5

f0005: 3D reconstructions of His-β-L HBcAg virus-like particles. (A) Cryomicroscopy of His-β-L HBcAg VLPs revealed two sizes 30 nm T = 3 and 34 nm T = 4. (B) The T = 3 structure was generated using 6048 particle images and achieved a resolution of 10 Å. (C) The T = 4 VLP was reconstructed using 2040 particles and achieved a resolution of 12 Å. Resolution assessment was by the FSC0.5 criterion (D). The general architecture of both capsids is similar to the wild type HBcAg, as demonstrated by comparison with a solvent exclusion surface representation of the T = 4 capsid calculated from the X-ray structure PDB ID 1QGT (E). His-β-L HBcAg particles however were seen to bear an extra spike density hypothesised to comprise a trimer of the extended N-terminus, located at sites of local threefold symmetry in the T = 3 VLP (magenta) and at both icosahedral (green) and local (magenta) threefold symmetry axes in the T = 4 particle. Wall-eyed stereo pair images are presented (B, C and E). Cross-sections through the T = 3 (F) and T = 4 (G) reconstructions show that the density in the N-terminal spikes is blurred compared to that of the HBcAg component (magenta and green arrows).
Mentions: Cryomicrographs of purified His-β-L HBcAg VLPs showed the presence of two populations of differently sized particles with diameters of approximately 30 nm and 34 nm, corresponding to the expected sizes for T = 3 and T = 4 classes (Fig. 1A). The T = 3 form was most common (74.4%), in line with the findings of previous studies showing that truncation of the C-terminus increases the ratio of T = 3 versus T = 4 particles (Pumpens and Grens, 2001). 7046 T = 3 and 2419 T = 4 particles were extracted from 785 micrographs and processed to calculate three-dimensional reconstructions.

Bottom Line: Heterologous sequences have been successfully inserted at both amino and carboxy termini as well as internally at the major immunodominant epitope.The insert was seen to form a trimeric spike on the capsid surface that was poorly resolved, most likely owing to it being flexible.We hypothesise that the capacity of N-terminal inserts to form trimers may have application in the development of multivalent vaccines to trimeric antigens.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Sir Michael Stoker Building, Garscube Campus, 464 Bearsden Road, Glasgow G61 1QH, Scotland, UK.

Show MeSH
Related in: MedlinePlus