Polymer Particles Bearing Recombinant LEL CD81 as Trapping Systems for Hepatitis C Virus

Hepatitis C is one of the most common social diseases in the world. The improvements in both the early diagnostics of the hepatitis C and the treatment of acute viremia caused by hepatitis C virus are undoubtedly an urgent task. In present work, we offered the micro- and nanotraps for the capturing of HCV. As a capturing moiety, we designed and synthesized in E. coli a fusion protein consisting of large extracellular loop of CD81 receptor and streptavidin as spacing part. The obtained protein has been immobilized on the surface of PLA-based micro- and nanoparticles. The developed trapping systems were characterized in terms of their physico-chemical properties. In order to illustrate the ability of developed micro- and nanotraps to bind HCV, E2 core protein of HCV was synthesized as a fusion protein with GFP. Interaction of E2 protein and hepatitis C virus-mimicking particles with the developed trapping systems were testified by several methods.

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Identification of the Hepatitis C Virus E2 Glycoprotein Binding Site on the Large Extracellular Loop of CD81

Journal of Virology ◽

10.1128/jvi.76.21.11143-11147.2002 ◽

2002 ◽

Vol 76 (21) ◽

pp. 11143-11147 ◽

Cited By ~ 97

Author(s):

Heidi E. Drummer ◽

Kirilee A. Wilson ◽

Pantelis Poumbourios

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Binding Site ◽

Nk Cell ◽

Cell Activity ◽

Random Mutagenesis ◽

Extracellular Loop ◽

Human T Cell ◽

Large Extracellular Loop

ABSTRACT The binding of hepatitis C virus glycoprotein E2 to the large extracellular loop (LEL) of CD81 has been shown to modulate human T-cell and NK cell activity in vitro. Using random mutagenesis of a chimera of maltose-binding protein and LEL residues 113 to 201, we have determined that the E2-binding site on CD81 comprises residues Ile182, Phe186, Asn184, and Leu162. These findings reveal an E2-binding surface of approximately 806 Å2 and potential target sites for the development of small-molecule inhibitors of E2 binding.

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Expression and Dimerization of Hepatitis C Virus Core Protein in E. coli

Hepatitis C Protocols ◽

10.1385/0-89603-521-2:325 ◽

2003 ◽

pp. 325-330 ◽

Cited By ~ 2

Author(s):

Shih-Yen Lo ◽

Jing-Hsiung Ou

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Core Protein ◽

E Coli ◽

Virus Core

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Expression of hepatitis C virus core protein as a fusion protein with maltose binding protein

Digestive Diseases and Sciences ◽

10.1007/bf01316791 ◽

1993 ◽

Vol 38 (4) ◽

pp. 626-630 ◽

Cited By ~ 3

Author(s):

Osamu Yokosuka ◽

Yoshimi Ito ◽

Jun Sakuma ◽

Fumio Imazeki ◽

Masao Ohto ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Fusion Protein ◽

Core Protein ◽

Binding Protein ◽

Maltose Binding Protein ◽

Virus Core

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Oligomerization of Hepatitis C Virus Core Protein Is Crucial for Interaction with the Cytoplasmic Domain of E1 Envelope Protein

Journal of Virology ◽

10.1128/jvi.01203-06 ◽

2006 ◽

Vol 80 (22) ◽

pp. 11265-11273 ◽

Cited By ~ 40

Author(s):

Kousuke Nakai ◽

Toru Okamoto ◽

Tomomi Kimura-Someya ◽

Koji Ishii ◽

Chang Kweng Lim ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Amino Acid ◽

Fusion Protein ◽

Core Protein ◽

Type I ◽

Amino Acid Residues ◽

Cytoplasmic Loop ◽

The Core ◽

Er Membrane

ABSTRACT Hepatitis C virus (HCV) contains two membrane-associated envelope glycoproteins, E1 and E2, which assemble as a heterodimer in the endoplasmic reticulum (ER). In this study, predictive algorithms and genetic analyses of deletion mutants and glycosylation site variants of the E1 glycoprotein were used to suggest that the glycoprotein can adopt two topologies in the ER membrane: the conventional type I membrane topology and a polytopic topology in which the protein spans the ER membrane twice with an intervening cytoplasmic loop (amino acid residues 288 to 360). We also demonstrate that the E1 glycoprotein is able to associate with the HCV core protein, but only upon oligomerization of the core protein in the presence of tRNA to form capsid-like structures. Yeast two-hybrid and immunoprecipitation analyses reveal that oligomerization of the core protein is promoted by amino acid residues 72 to 91 in the core. Furthermore, the association between the E1 glycoprotein and the assembled core can be recapitulated using a fusion protein containing the putative cytoplasmic loop of the E1 glycoprotein. This fusion protein is also able to compete with the intact E1 glycoprotein for binding to the core. Mutagenesis of the cytoplasmic loop of E1 was used to define a region of four amino acids (residues 312 to 315) that is important for interaction with the assembled HCV core. Taken together, our studies suggest that interaction between the self-oligomerized HCV core and the E1 glycoprotein is mediated through the cytoplasmic loop present in a polytopic form of the E1 glycoprotein.

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CD81 Receptor Regions outside the Large Extracellular Loop Determine Hepatitis C Virus Entry into Hepatoma Cells

Viruses ◽

10.3390/v10040207 ◽

2018 ◽

Vol 10 (4) ◽

pp. 207 ◽

Cited By ~ 7

Author(s):

Pia Banse ◽

Rebecca Moeller ◽

Janina Bruening ◽

Lisa Lasswitz ◽

Sina Kahl ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Virus Entry ◽

Hepatoma Cells ◽

Extracellular Loop ◽

Large Extracellular Loop

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Disulfide Bonds in Hepatitis C Virus Glycoprotein E1 Control the Assembly and Entry Functions of E2 Glycoprotein

Journal of Virology ◽

10.1128/jvi.02659-12 ◽

2012 ◽

Vol 87 (3) ◽

pp. 1605-1617 ◽

Cited By ~ 35

Author(s):

Ahmed Wahid ◽

François Helle ◽

Véronique Descamps ◽

Gilles Duverlie ◽

François Penin ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Fusion Protein ◽

Membrane Fusion ◽

Reverse Genetics ◽

Core Protein ◽

Binding Capacity ◽

Class Ii ◽

Wild Type Virus ◽

Additional Function

ABSTRACTClass II membrane fusion proteins have been described in viruses in which the envelope proteins are derived from a precursor polyprotein containing two transmembrane glycoproteins arranged in tandem. Although the second protein, which carries the membrane fusion function, is in general well characterized, the companion protein, which is a protein chaperone for the folding of the fusion protein, is less well characterized for some viruses, like hepatitis C virus (HCV). To investigate the role of the class II companion glycoprotein E1 of HCV, we chose to target conserved cysteine residues in the protein, and we systematically mutated them in a full-length infectious HCV clone by reverse genetics. All the mutants were infectious, albeit with lower titers than the wild-type virus. The reduced infectivity was in part due to a decrease in viral assembly, as revealed by measurement of intracellular infectivity and by quantification of core protein released from cells transfected with mutant genomes. Analyses of mutated proteins did not show any major defect in folding. However, the mutations reduced virus stability, and they could also affect the density of infectious viral particles. Mutant viruses also showed a defect in cell-to-cell transmission. Finally, our data indicate that HCV glycoprotein E1 can also affect the fusion protein E2 by modulating its recognition by the cellular coreceptor CD81. Therefore, in the context of HCV, our data identify an additional function of a class II companion protein as a molecule that can control the binding capacity of the fusion protein.

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Mechanism of Structural Tuning of the Hepatitis C Virus Human Cellular Receptor CD81 Large Extracellular Loop

Structure ◽

10.1016/j.str.2017.11.015 ◽

2018 ◽

Vol 26 (1) ◽

pp. 181 ◽

Cited By ~ 1

Author(s):

Eva S. Cunha ◽

Pedro Sfriso ◽

Adriana L. Rojas ◽

Pietro Roversi ◽

Adam Hospital ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Cellular Receptor ◽

Extracellular Loop ◽

Large Extracellular Loop

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Diverse CD81 Proteins Support Hepatitis C Virus Infection

Journal of Virology ◽

10.1128/jvi.00104-06 ◽

2006 ◽

Vol 80 (22) ◽

pp. 11331-11342 ◽

Cited By ~ 117

Author(s):

Mike Flint ◽

Thomas von Hahn ◽

Jie Zhang ◽

Michelle Farquhar ◽

Christopher T. Jones ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Amino Acid ◽

Viral Entry ◽

Hepatitis C Virus Infection ◽

Extracellular Loop ◽

Virus Attachment ◽

Susceptibility To Infection ◽

Cell Incubation ◽

Large Extracellular Loop

ABSTRACT Hepatitis C virus (HCV) entry is dependent on CD81. To investigate whether the CD81 sequence is a determinant of HCV host range, we expressed a panel of diverse CD81 proteins and tested their ability to interact with HCV. CD81 large extracellular loop (LEL) sequences were expressed as recombinant proteins; the human and, to a low level, the African green monkey sequences bound soluble HCV E2 (sE2) and inhibited infection by retrovirus pseudotype particles bearing HCV glycoproteins (HCVpp). In contrast, mouse or rat CD81 proteins failed to bind sE2 or to inhibit HCVpp infection. However, CD81 proteins from all species, when expressed in HepG2 cells, conferred susceptibility to infection by HCVpp and cell culture-grown HCV to various levels, with the rat sequence being the least efficient. Recombinant human CD81 LEL inhibited HCVpp infectivity only if present during the virus-cell incubation, consistent with a role for CD81 after virus attachment. Amino acid changes that abrogate sE2 binding (I182F, N184Y, and F186S, alone or in combination) were introduced into human CD81. All three amino acid changes in human CD81 resulted in a molecule that still supported HCVpp infection, albeit with reduced efficiency. In summary, there is a remarkable plasticity in the range of CD81 sequences that can support HCV entry, suggesting that CD81 polymorphism may contribute to, but alone does not define, the HCV susceptibility of a species. In addition, the capacity to support viral entry is only partially reflected by assays measuring sE2 interaction with recombinant or full-length CD81 proteins.

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