Hepatitis C virus E2 protein purified from mammalian cells is frequently recognized by E2-specific antibodies in patient sera.
The envelope protein of hepatitis C virus (HCV) is composed of two membrane-associated glycoproteins, E1 and E2. To obtain HCV E2 protein as a secretory form at a high level, we constructed a recombinant chinese hamster ovary (CHO) cell line expressing a C-terminal truncated E2 (E2t) fused to human growth hormone (hGH), CHO/hGHE2t. The hGHE2t fusion protein was purified from the culture supernatant using anti-hGH mAb affinity chromatography at approximately 80% purity. The purified hGHE2t protein appeared to be assembled into oligomers linked by intermolecular disulfide bond(s) when density gradient centrifugation and SDS-polyacrylamide gel electrophoresis were employed. When the purified fusion protein was used for testing its ability to bind to antibodies specific for HCV by enzyme-linked immunosorbent assay, the protein was recognized by antibodies in sera from 90% of HCV-positive patients. Treatment of hGHE2t protein by beta-mercaptoethanol, but not by heat and SDS, significantly reduced its reactivity to the antibodies of patient sera, suggesting that intermolecular and/or intramolecular disulfide bonds are important for its ability to recognize its specific antibody and that the E2 protein contains discontinuous antigenic epitope(s).
Lee
K J
KJ
Department of Life Science, Center for Biofunctional Molecules, School of Environmental Engineering, Pohang University of Science and Technology, San 31, Hyoja Dong, Pohang, 790-784, Korea.
Suh
Y A
YA
Cho
Y G
YG
Cho
Y S
YS
Ha
G W
GW
Chung
K H
KH
Hwang
J H
JH
Yun
Y D
YD
Lee
D S
DS
Kim
C M
CM
Sung
Y C
YC
D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology*; D016466:CHO Cells; D006224:Cricetinae; D004220:Disulfides / Q000737:chemistry; D016174:Hepacivirus / Q000276:immunology*; D018963:Hepatitis Antigens / Q000276:immunology*; D006526:Hepatitis C / Q000276:immunology*; D006801:Humans; D011489:Protein Denaturation; D011993:Recombinant Fusion Proteins / Q000276:immunology; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism
D000914:Antibodies, Viral; D004220:Disulfides; D018963:Hepatitis Antigens; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus