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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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ResMap analysis of His-β-L HBcAg VLP reconstructions. ResMap was used to evaluate the local resolution of both T = 3 (A) and T = 4 (B) reconstructions. Isosurfaced representations of the reconstructions were coloured accordingly revealing that the HBcAg component was solved at higher resolution than the N-terminal extension. In the case of the T = 3 particle the core density achieved resolution approaching 5 Å, ranging to 8 Å in places, while the inserted polypeptide was seen to be poorer than 19 Å resolution. The T = 4 reconstruction was calculated from fewer particles and achieved lower resolution ranging between 8 and 10 Å in the HBcAg region and 11–>19 Å in the N-terminal extension. Interestingly both ResMap analyses gave resolution assessments that included small patches of poor-resolution throughout the structure, in particular within the dimeric spike of HBcAg, these are apparent in clipped isosurface representations (C and D).
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f0010: ResMap analysis of His-β-L HBcAg VLP reconstructions. ResMap was used to evaluate the local resolution of both T = 3 (A) and T = 4 (B) reconstructions. Isosurfaced representations of the reconstructions were coloured accordingly revealing that the HBcAg component was solved at higher resolution than the N-terminal extension. In the case of the T = 3 particle the core density achieved resolution approaching 5 Å, ranging to 8 Å in places, while the inserted polypeptide was seen to be poorer than 19 Å resolution. The T = 4 reconstruction was calculated from fewer particles and achieved lower resolution ranging between 8 and 10 Å in the HBcAg region and 11–>19 Å in the N-terminal extension. Interestingly both ResMap analyses gave resolution assessments that included small patches of poor-resolution throughout the structure, in particular within the dimeric spike of HBcAg, these are apparent in clipped isosurface representations (C and D).

Mentions: Resmap analysis (Kucukelbir et al., 2014) of the T = 3 reconstruction indicated a mean resolution of 7.8 Å; significantly better than the estimate derived by the FSC0.5 method. Analysis of the T = 4 structure also indicated a better resolution than that estimated by Fourier shell correlation at 0.5 cutoff, the mean resolution was determined to be 10.6 Å (Fig. 2). Local resolution measurements were seen to range between 5 and 8 Å within the core regions of the T = 3 VLP, while in the T = 4 structure the HBcAg components were resolved at between 8 and 10 Å. Interestingly in both analyses the novel density that we attribute to the N-terminal extension was highlighted as having a substantially lower resolution than the HBcAg structure. For the T = 3 particle this region was determined to have a resolution poorer than 19 Å, likewise in the T = 4 structure the trimeric spike was identified as a region of low resolution.


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)

ResMap analysis of His-β-L HBcAg VLP reconstructions. ResMap was used to evaluate the local resolution of both T = 3 (A) and T = 4 (B) reconstructions. Isosurfaced representations of the reconstructions were coloured accordingly revealing that the HBcAg component was solved at higher resolution than the N-terminal extension. In the case of the T = 3 particle the core density achieved resolution approaching 5 Å, ranging to 8 Å in places, while the inserted polypeptide was seen to be poorer than 19 Å resolution. The T = 4 reconstruction was calculated from fewer particles and achieved lower resolution ranging between 8 and 10 Å in the HBcAg region and 11–>19 Å in the N-terminal extension. Interestingly both ResMap analyses gave resolution assessments that included small patches of poor-resolution throughout the structure, in particular within the dimeric spike of HBcAg, these are apparent in clipped isosurface representations (C and D).
© Copyright Policy - CC BY
Related In: Results  -  Collection

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

f0010: ResMap analysis of His-β-L HBcAg VLP reconstructions. ResMap was used to evaluate the local resolution of both T = 3 (A) and T = 4 (B) reconstructions. Isosurfaced representations of the reconstructions were coloured accordingly revealing that the HBcAg component was solved at higher resolution than the N-terminal extension. In the case of the T = 3 particle the core density achieved resolution approaching 5 Å, ranging to 8 Å in places, while the inserted polypeptide was seen to be poorer than 19 Å resolution. The T = 4 reconstruction was calculated from fewer particles and achieved lower resolution ranging between 8 and 10 Å in the HBcAg region and 11–>19 Å in the N-terminal extension. Interestingly both ResMap analyses gave resolution assessments that included small patches of poor-resolution throughout the structure, in particular within the dimeric spike of HBcAg, these are apparent in clipped isosurface representations (C and D).
Mentions: Resmap analysis (Kucukelbir et al., 2014) of the T = 3 reconstruction indicated a mean resolution of 7.8 Å; significantly better than the estimate derived by the FSC0.5 method. Analysis of the T = 4 structure also indicated a better resolution than that estimated by Fourier shell correlation at 0.5 cutoff, the mean resolution was determined to be 10.6 Å (Fig. 2). Local resolution measurements were seen to range between 5 and 8 Å within the core regions of the T = 3 VLP, while in the T = 4 structure the HBcAg components were resolved at between 8 and 10 Å. Interestingly in both analyses the novel density that we attribute to the N-terminal extension was highlighted as having a substantially lower resolution than the HBcAg structure. For the T = 3 particle this region was determined to have a resolution poorer than 19 Å, likewise in the T = 4 structure the trimeric spike was identified as a region of low resolution.

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