Expression and structural properties of a chimeric protein based on the ectodomains of E1 and E2 hepatitis C virus envelope glycoproteins.
Hepatitis C virus encodes two enveloped glycoproteins, E1 and E2, which are involved in viral attachment and entry into target cells. We have obtained in insect cells infected by recombinant baculovirus a chimeric secreted recombinant protein, E1(341)E2(661,) containing the ectodomains of E1 and E2. The described procedure allows the purification of approximately 2mg of protein from 1L of culture media. Sedimentation velocity experiments and SDS-PAGE in the absence of reducing agents indicate that the protein has a high tendency to self-associate, the dimer being the main species observed. All the oligomeric forms observed maintain a conformation which is recognized by the conformation-dependent monoclonal antibody H53 directed against the E2 ectodomain. The spectroscopic properties of E1(341)E2(661) are those of a three-dimensionally structured protein. Moreover, the chimeric protein is able to bind to human antibodies present in HCV-positive human sera. Accordingly, this chimeric soluble polypeptide chain may be a valuable tool to study the structure-function relationship of HCV envelope proteins.
Tello
Daniel
D
Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Químicas, Universidad Complutense, Madrid, Spain.
Rodríguez-Rodríguez
Mar
M
Yélamos
Belén
B
Gómez-Gutiérrez
Julián
J
Ortega
Sara
S
Pacheco
Beatriz
B
Peterson
Darrell L
DL
Gavilanes
Francisco
F
D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D016367:Baculoviridae / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D016174:Hepacivirus / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D006801:Humans; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000378:metabolism; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus