Intracellular Expression of a Cloned Antibody Fragment Interferes with Hepatitis B Virus Surface Antigen Secretion

We evaluated the potential of an intracellularly expressed antibody fragment to interfere with hepatitis B virus (HBV). Sequences coding for the immunoglobulin variable regions of the HBV surface antigen (HBsAg) specific monoclonal antibody 5C3 were isolated and characterized. A secretory pathway-ta...

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Bibliographic Details
Published in:Biochemical and biophysical research communications 1999-02, Vol.255 (3), p.785-791
Main Authors: Putlitz, Jasper zu, Skerra, Arne, Wands, Jack R.
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Base Sequence
Carcinoma, Hepatocellular - immunology
Cloning, Molecular
Fluorescent Antibody Technique
Hepatitis B Surface Antigens - immunology
Hepatitis B virus
Hepatitis Viruses - immunology
Humans
Immunoglobulin Fragments - genetics
Immunoglobulin Fragments - immunology
Microscopy, Confocal
Molecular Sequence Data
Protein Sorting Signals - immunology
Sequence Analysis, DNA
Transfection - genetics
Tumor Cells, Cultured
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Summary:We evaluated the potential of an intracellularly expressed antibody fragment to interfere with hepatitis B virus (HBV). Sequences coding for the immunoglobulin variable regions of the HBV surface antigen (HBsAg) specific monoclonal antibody 5C3 were isolated and characterized. A secretory pathway-targeted, 5C3 derived single chain Fv (sFv) fragment was expressed in HuH-7 hepatocellular carcinoma cells together with HBsAg. Quantification of extracellular HBsAg levels in the cell culture supernatant demonstrated that the presence of the 5C3 sFv equipped with a secretory pathway retention signal SEKDEL reduced extracellular HBsAg levels by a mean of 85%. Co-immunoprecipitation studies revealed that the 5C3 sFv targeted to the secretory pathway physically interacted with its target antigen, HBsAg. Confocal microscopy studies confirmed the intracellular expression and colocalization of the 5C3 sFv and HBsAg. We conclude that certain intracellularly expressed antibody fragments will substantially interfere with HBV antigen secretion from the cell.
ISSN:0006-291X
1090-2104
DOI:10.1006/bbrc.1999.0149