Structural characterisation of the hepatitis C envelope glycoprotein E1 ectodomain derived from a mammalian and a yeast expression system

Lorent, Eric; Bierau, Horst; Engelborghs, Yves; Verheyden, Gert; Bosman, Fons

doi:10.1016/j.vaccine.2007.11.004

Cited by 11 publications

(20 citation statements)

References 42 publications

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“…Highly purified recombinant HCV E1 191‐326 (Innogenetics Biologicals, Gent, Belgium)27 and E2 384‐715 (kindly provided by Michael Houghton; Chiron Corporation, Emeryville, CA) were examined for binding with ApoB and ApoE (Calbiochem). Briefly, an ELISA plate (Maxisorp; Nunc Corporation, Rochester, NY) was coated with ApoB or ApoE (100 ng/well) in PBS (pH 7.4) overnight at 4°C.…”

Section: Methodsmentioning

confidence: 99%

Hepatitis C virus E1 envelope glycoprotein interacts with apolipoproteins in facilitating entry into hepatocytes

et al. 2011

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Our previous studies demonstrated that hepatitis C virus (HCV) envelope glycoproteins, E1 and E2, display distinct reactivity to different cell surface molecules. In this study, we characterized the interaction of E1 and E2 with apolipoproteins in facilitating virus entry. The results suggested a higher neutralization of VSV/HCV E1-G pseudotype infectivity by antibodies to apolipoprotein E (ApoE) than apolipoprotein B (ApoB), with VSV/HCV E2-G pseudotype infectivity remaining largely unaffected. Neutralization of cell culture grown HCV infectivity by antiserum to ApoE, and to a lesser extent by ApoB, further verified their involvement in virus entry. HCV E1, but not E2, displayed binding with ApoE and ApoB by ELISA. Binding of E1 with apolipoproteins were further supported by coimmunoprecipitation from human hepatocytes expressing E1. Rabbit antiserum to a selected E1 ectodomain derived peptide displayed ~50% neutralization of E1-G pseudotype infectivity. Furthermore, E1 ectodomain derived synthetic peptides significantly inhibited the interaction of E1 with both the apolipoproteins. Investigation on the role of LDL-R as a hepatocyte surface receptor for virus entry suggested a significant reduction in E1-G pseudotype plaque numbers (~70%) by inhibiting LDL-R ligand binding activity using human proprotein convertase subtilisin/kexin type 9 (PCSK9) and platelet factor-4 (PF4), while they had a minimal inhibitory effect on E2-G pseudotype. Conclusion Together, the results suggested an association between HCV E1 and apolipoproteins, which may facilitate virus entry through LDL-R into mammalian cells.

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Section: Methodsmentioning

confidence: 99%

Hepatitis C virus E1 envelope glycoprotein interacts with apolipoproteins in facilitating entry into hepatocytes

et al. 2011

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“…The cellular mechanism of HCV entry has been studied using HCV pseudoparticles (HCVpp), infectious retroviral particles with HCV envelope proteins on the surface, and the cell culture model which allows for the production and propagation of virus in cell culture (HCVcc) (Lindenbach and Rice, 2005;Wakita et al, 2005;Zhong et al, 2005). Despite that E1 and E2 have been expressed in several prokaryotic (Hüssy et al, 1997) or eukaryotic (Lorent et al, 2008;Mustilli et al, 1999;Michalak et al, 1997;Hüssy et al, 1996) cell lines, few data concerning to the structure of isolated proteins have been obtained. Most of the structural studies have been focused on the E2 protein, since, contrary to the full-length polypeptide, the E2 ectodomain has been characterized as an independent folding unit (Rodriguez-Rodriguez et al, 2009;Whidby et al, 2009).…”

Section: Introductionmentioning

confidence: 99%

High-yield production of a chimeric glycoprotein based on permuted E1 and E2 HCV envelope ectodomains

Tello

Rodríguez-Rodríguez

Yelamos

et al. 2015

Journal of Virological Methods

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In this report it is described for the first time the expression and purification of large quantities of a soluble and correctly folded chimeric recombinant protein, E2 661 E1 340, containing the permuted Hepatitis C virus (HCV) glycoprotein ectodomains E1 (aminoacids 192-340) and E2 (aminoacids 384-661). Using the baculovirus/insect cell expression system, 8 mg of secreted protein were purified from 1 L of culture media, a yield 4 times higher than the described for its counterpart E1 341 E2 661 . This permuted chimeric protein is glycosylated and possesses a high tendency to selfassociate. The fluorescence emission spectrum indicates that Trp residues occupy a relatively low hydrophobic environment. The secondary structure was determined by deconvolution of the far-UV circular dichroism spectrum yielding 13% -helix structure, 49% extended structure and 38% non-ordered structure. E2 661 E1 340 binds to antibodies present in human sera from HCV-positive patients, a binding that is blocked at different levels by a rabbit anti-E2 661 antibody. All these structural and antigenic features of E2 661 E1 340 are very similar to those described for E1 340 E2 661 , Thus, this high-yield isolated chimeric protein may be a valuable tool to study the first steps of the HCV infection.

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“…Consequently, the development of vaccines against HCV infection has been markedly stalled by the complexity and heterogeneity of the virus. To date, there has not been an efficient vaccine clinically validated 13…”

Section: Introductionmentioning

confidence: 99%

“…E1 from both sources was shown to spontaneously form protein particles in the absence of detergents. Detergents cause the particles to dissociate into monomers 13. The secondary and tertiary structure of the E1 monomers and particles differed significantly in their circular dichroism and intrinsic (Trp) fluorescence but was similar as produced by the two expression systems 13.…”

Section: Introductionmentioning

confidence: 99%

“…Detergents cause the particles to dissociate into monomers 13. The secondary and tertiary structure of the E1 monomers and particles differed significantly in their circular dichroism and intrinsic (Trp) fluorescence but was similar as produced by the two expression systems 13. Despite small variations in tryptophan fluorescence, E1 particles produced by yeast expression displayed similar structural and immunogenetic properties compared to those derived from mammalian cells 13…”

Section: Introductionmentioning

confidence: 99%

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Structural stability of hepatitis C virus envelope glycoprotein E1: Effect of pH and dissociative detergents

Joshi

Bosman

et al. 2009

Journal of Pharmaceutical Sciences

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Structural characterisation of the hepatitis C envelope glycoprotein E1 ectodomain derived from a mammalian and a yeast expression system

Cited by 11 publications

References 42 publications

Hepatitis C virus E1 envelope glycoprotein interacts with apolipoproteins in facilitating entry into hepatocytes

Hepatitis C virus E1 envelope glycoprotein interacts with apolipoproteins in facilitating entry into hepatocytes

High-yield production of a chimeric glycoprotein based on permuted E1 and E2 HCV envelope ectodomains

Structural stability of hepatitis C virus envelope glycoprotein E1: Effect of pH and dissociative detergents

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