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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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Homology modelling of His-β-L HBcAg. To evaluate the structure of the N-terminal extension of His-β-L HBcAg the sequence was homology modelled in MOE against the known structure of HBcAg with outgap modelling to predict the structure of the extension. The models revealed a largely unstructured polypeptide with regions of α-helix close to the N-terminus. The best model is shown (rainbow – blue = N terminus, red = C terminus) as a stereo pair viewed from the top (A) and side (B) and as a dimer with the HBcAg structure (grey) for comparison. In this model the N-terminus is oriented towards the capsid interior. Most likely this is not correct as this region includes the polyhistidine tract that was employed in purifying the particles by immobilised metal affinity chromatography. Moreover docking the model into our sharpened T = 3 reconstruction (thresholded to enclose the correct molecular volume) reveals that the model does not fit into the reconstructed density and there is substantial collision with symmetry related protomers (C and D). Three dimers are shown arranged about the N-terminal spike feature at a local threefold symmetry axis. Each monomer that contributes an N-terminal extension to the spike feature is shown in the rainbow colour scheme, while the symmetry related monomers are shown as the HBcAg structure in grey.
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f0025: Homology modelling of His-β-L HBcAg. To evaluate the structure of the N-terminal extension of His-β-L HBcAg the sequence was homology modelled in MOE against the known structure of HBcAg with outgap modelling to predict the structure of the extension. The models revealed a largely unstructured polypeptide with regions of α-helix close to the N-terminus. The best model is shown (rainbow – blue = N terminus, red = C terminus) as a stereo pair viewed from the top (A) and side (B) and as a dimer with the HBcAg structure (grey) for comparison. In this model the N-terminus is oriented towards the capsid interior. Most likely this is not correct as this region includes the polyhistidine tract that was employed in purifying the particles by immobilised metal affinity chromatography. Moreover docking the model into our sharpened T = 3 reconstruction (thresholded to enclose the correct molecular volume) reveals that the model does not fit into the reconstructed density and there is substantial collision with symmetry related protomers (C and D). Three dimers are shown arranged about the N-terminal spike feature at a local threefold symmetry axis. Each monomer that contributes an N-terminal extension to the spike feature is shown in the rainbow colour scheme, while the symmetry related monomers are shown as the HBcAg structure in grey.

Mentions: To further understand the structure of the N-terminal extension in His-β-L HBcAg VLPs we used MOE to model the tertiary structure of the protein. The models judged to be the best predicted that the extension would contain an unstructured region followed by an alpha helix located between residues Met12-Asp28, Ser4-Asp24 or His10-Asp28 in the top 3 models. Thus in two models the alpha helix was C-terminal to the polyhistidine tract while the remaining model incorporated the 6HIS motif into the helix. The length of the alpha helix varied in the different models ranging between 24 and 29 Å. Upon comparison with our T = 3 structure the models did not match the reconstructed density well, the orientation of the N-terminal extension varied widely between predicted structures and in all cases clashed with symmetry related protomers (Fig. 5). As the reconstructed density for the N-terminal extension was poorly resolved, no attempt was made to refine the models by flexible fitting.


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)

Homology modelling of His-β-L HBcAg. To evaluate the structure of the N-terminal extension of His-β-L HBcAg the sequence was homology modelled in MOE against the known structure of HBcAg with outgap modelling to predict the structure of the extension. The models revealed a largely unstructured polypeptide with regions of α-helix close to the N-terminus. The best model is shown (rainbow – blue = N terminus, red = C terminus) as a stereo pair viewed from the top (A) and side (B) and as a dimer with the HBcAg structure (grey) for comparison. In this model the N-terminus is oriented towards the capsid interior. Most likely this is not correct as this region includes the polyhistidine tract that was employed in purifying the particles by immobilised metal affinity chromatography. Moreover docking the model into our sharpened T = 3 reconstruction (thresholded to enclose the correct molecular volume) reveals that the model does not fit into the reconstructed density and there is substantial collision with symmetry related protomers (C and D). Three dimers are shown arranged about the N-terminal spike feature at a local threefold symmetry axis. Each monomer that contributes an N-terminal extension to the spike feature is shown in the rainbow colour scheme, while the symmetry related monomers are shown as the HBcAg structure in grey.
© Copyright Policy - CC BY
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4318616&req=5

f0025: Homology modelling of His-β-L HBcAg. To evaluate the structure of the N-terminal extension of His-β-L HBcAg the sequence was homology modelled in MOE against the known structure of HBcAg with outgap modelling to predict the structure of the extension. The models revealed a largely unstructured polypeptide with regions of α-helix close to the N-terminus. The best model is shown (rainbow – blue = N terminus, red = C terminus) as a stereo pair viewed from the top (A) and side (B) and as a dimer with the HBcAg structure (grey) for comparison. In this model the N-terminus is oriented towards the capsid interior. Most likely this is not correct as this region includes the polyhistidine tract that was employed in purifying the particles by immobilised metal affinity chromatography. Moreover docking the model into our sharpened T = 3 reconstruction (thresholded to enclose the correct molecular volume) reveals that the model does not fit into the reconstructed density and there is substantial collision with symmetry related protomers (C and D). Three dimers are shown arranged about the N-terminal spike feature at a local threefold symmetry axis. Each monomer that contributes an N-terminal extension to the spike feature is shown in the rainbow colour scheme, while the symmetry related monomers are shown as the HBcAg structure in grey.
Mentions: To further understand the structure of the N-terminal extension in His-β-L HBcAg VLPs we used MOE to model the tertiary structure of the protein. The models judged to be the best predicted that the extension would contain an unstructured region followed by an alpha helix located between residues Met12-Asp28, Ser4-Asp24 or His10-Asp28 in the top 3 models. Thus in two models the alpha helix was C-terminal to the polyhistidine tract while the remaining model incorporated the 6HIS motif into the helix. The length of the alpha helix varied in the different models ranging between 24 and 29 Å. Upon comparison with our T = 3 structure the models did not match the reconstructed density well, the orientation of the N-terminal extension varied widely between predicted structures and in all cases clashed with symmetry related protomers (Fig. 5). As the reconstructed density for the N-terminal extension was poorly resolved, no attempt was made to refine the models by flexible fitting.

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