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Structure of the core ectodomain of the hepatitis C virus envelope glycoprotein 2.

Khan AG, Whidby J, Miller MT, Scarborough H, Zatorski AV, Cygan A, Price AA, Yost SA, Bohannon CD, Jacob J, Grakoui A, Marcotrigiano J - Nature (2014)

Bottom Line: Sheet A has an IgG-like fold that is commonly found in viral and cellular proteins, whereas sheet B represents a novel fold.Solution-based studies demonstrate that the full-length E2 ectodomain has a similar globular architecture and does not undergo significant conformational or oligomeric rearrangements on exposure to low pH.These results provide unprecedented insights into HCV entry and will assist in developing an HCV vaccine and new inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, 679 Hoes Lane West, Piscataway, New Jersey 08854, USA.

ABSTRACT
Hepatitis C virus (HCV) is a significant public health concern with approximately 160 million people infected worldwide. HCV infection often results in chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. No vaccine is available and current therapies are effective against some, but not all, genotypes. HCV is an enveloped virus with two surface glycoproteins (E1 and E2). E2 binds to the host cell through interactions with scavenger receptor class B type I (SR-BI) and CD81, and serves as a target for neutralizing antibodies. Little is known about the molecular mechanism that mediates cell entry and membrane fusion, although E2 is predicted to be a class II viral fusion protein. Here we describe the structure of the E2 core domain in complex with an antigen-binding fragment (Fab) at 2.4 Å resolution. The E2 core has a compact, globular domain structure, consisting mostly of β-strands and random coil with two small α-helices. The strands are arranged in two, perpendicular sheets (A and B), which are held together by an extensive hydrophobic core and disulphide bonds. Sheet A has an IgG-like fold that is commonly found in viral and cellular proteins, whereas sheet B represents a novel fold. Solution-based studies demonstrate that the full-length E2 ectodomain has a similar globular architecture and does not undergo significant conformational or oligomeric rearrangements on exposure to low pH. Thus, the IgG-like fold is the only feature that E2 shares with class II membrane fusion proteins. These results provide unprecedented insights into HCV entry and will assist in developing an HCV vaccine and new inhibitors.

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E2 core contains an extensive hydrophobic coreSheets A and B are held together by an extensive hydrophobic core composed of mostly aromatic amino acids (green) and five disulfide bonds (yellow).
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Figure 5: E2 core contains an extensive hydrophobic coreSheets A and B are held together by an extensive hydrophobic core composed of mostly aromatic amino acids (green) and five disulfide bonds (yellow).

Mentions: Monoclonal antibodies were generated against recombinant eE2 and crystals of deglycosylated E2 core were produced in complex with an Fab (2A12) to 2.4 Å resolution (Fig. 1 and Extended Data Table 1). The complex structure was determined by molecular replacement using an Fab structure followed by iterative rounds of model building and refinement. The E2 core domain has a globular fold, consisting of mostly beta strands and random coil with two short alpha helices, which is consistent with previous spectroscopic studies of eE2 12,13. The protein contains two, four-stranded antiparallel beta sheets (termed sheets A and B), the planes of which are approximately perpendicular to each other. The four strands of the amino-terminal beta sheet (sheet A) are stabilized by two disulfide bonds, between strands 1 and 3 [C7 (510) and C8 (554)] and the amino terminal loop with strand 4 [C5 (496) and C9 (566)]. The loop between strands 2 and 3 contains sequences implicated in CD81 binding 14,15 and is flexible, similar to the amino-terminal CD81 binding sites, which were deleted. After strand 4, the polypeptide continues into a long, disordered loop before forming the first short helix (H1) followed by the second beta sheet (sheet B). A second short alpha helix (H2) is located between strands 6 and 7. A disulfide bond [C14 (611) and C16 (648)] between strand 6 and the carboxyl-terminal strand 8 further stabilizes the fold. The carboxyl-terminal strands (7 and 8) are the longest within the protein with approximately nine amino acids each and encompass the 2A12-binding site. 2A12 does not neutralize HCV infection, suggesting that the epitope is either buried within the particle or incapable of preventing entry (Extended Data Fig. 4f). The two beta sheets are held together by i) two disulfide bonds, connecting the loops before strand 1 and after H2 [C4 (488) with C15 (624)] as well as the loops after strand 4 and before H1 [C10 (571) and C13 (601)], and ii) an extensive hydrophobic core consisting of numerous aromatic residues (Extended Data Fig. 5).


Structure of the core ectodomain of the hepatitis C virus envelope glycoprotein 2.

Khan AG, Whidby J, Miller MT, Scarborough H, Zatorski AV, Cygan A, Price AA, Yost SA, Bohannon CD, Jacob J, Grakoui A, Marcotrigiano J - Nature (2014)

E2 core contains an extensive hydrophobic coreSheets A and B are held together by an extensive hydrophobic core composed of mostly aromatic amino acids (green) and five disulfide bonds (yellow).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4126800&req=5

Figure 5: E2 core contains an extensive hydrophobic coreSheets A and B are held together by an extensive hydrophobic core composed of mostly aromatic amino acids (green) and five disulfide bonds (yellow).
Mentions: Monoclonal antibodies were generated against recombinant eE2 and crystals of deglycosylated E2 core were produced in complex with an Fab (2A12) to 2.4 Å resolution (Fig. 1 and Extended Data Table 1). The complex structure was determined by molecular replacement using an Fab structure followed by iterative rounds of model building and refinement. The E2 core domain has a globular fold, consisting of mostly beta strands and random coil with two short alpha helices, which is consistent with previous spectroscopic studies of eE2 12,13. The protein contains two, four-stranded antiparallel beta sheets (termed sheets A and B), the planes of which are approximately perpendicular to each other. The four strands of the amino-terminal beta sheet (sheet A) are stabilized by two disulfide bonds, between strands 1 and 3 [C7 (510) and C8 (554)] and the amino terminal loop with strand 4 [C5 (496) and C9 (566)]. The loop between strands 2 and 3 contains sequences implicated in CD81 binding 14,15 and is flexible, similar to the amino-terminal CD81 binding sites, which were deleted. After strand 4, the polypeptide continues into a long, disordered loop before forming the first short helix (H1) followed by the second beta sheet (sheet B). A second short alpha helix (H2) is located between strands 6 and 7. A disulfide bond [C14 (611) and C16 (648)] between strand 6 and the carboxyl-terminal strand 8 further stabilizes the fold. The carboxyl-terminal strands (7 and 8) are the longest within the protein with approximately nine amino acids each and encompass the 2A12-binding site. 2A12 does not neutralize HCV infection, suggesting that the epitope is either buried within the particle or incapable of preventing entry (Extended Data Fig. 4f). The two beta sheets are held together by i) two disulfide bonds, connecting the loops before strand 1 and after H2 [C4 (488) with C15 (624)] as well as the loops after strand 4 and before H1 [C10 (571) and C13 (601)], and ii) an extensive hydrophobic core consisting of numerous aromatic residues (Extended Data Fig. 5).

Bottom Line: Sheet A has an IgG-like fold that is commonly found in viral and cellular proteins, whereas sheet B represents a novel fold.Solution-based studies demonstrate that the full-length E2 ectodomain has a similar globular architecture and does not undergo significant conformational or oligomeric rearrangements on exposure to low pH.These results provide unprecedented insights into HCV entry and will assist in developing an HCV vaccine and new inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, 679 Hoes Lane West, Piscataway, New Jersey 08854, USA.

ABSTRACT
Hepatitis C virus (HCV) is a significant public health concern with approximately 160 million people infected worldwide. HCV infection often results in chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. No vaccine is available and current therapies are effective against some, but not all, genotypes. HCV is an enveloped virus with two surface glycoproteins (E1 and E2). E2 binds to the host cell through interactions with scavenger receptor class B type I (SR-BI) and CD81, and serves as a target for neutralizing antibodies. Little is known about the molecular mechanism that mediates cell entry and membrane fusion, although E2 is predicted to be a class II viral fusion protein. Here we describe the structure of the E2 core domain in complex with an antigen-binding fragment (Fab) at 2.4 Å resolution. The E2 core has a compact, globular domain structure, consisting mostly of β-strands and random coil with two small α-helices. The strands are arranged in two, perpendicular sheets (A and B), which are held together by an extensive hydrophobic core and disulphide bonds. Sheet A has an IgG-like fold that is commonly found in viral and cellular proteins, whereas sheet B represents a novel fold. Solution-based studies demonstrate that the full-length E2 ectodomain has a similar globular architecture and does not undergo significant conformational or oligomeric rearrangements on exposure to low pH. Thus, the IgG-like fold is the only feature that E2 shares with class II membrane fusion proteins. These results provide unprecedented insights into HCV entry and will assist in developing an HCV vaccine and new inhibitors.

Show MeSH
Related in: MedlinePlus