Fine mapping of neutralization epitopes on duck hepatitis B virus (DHBV) pre-S protein using monoclonal antibodies and overlapping peptides.
To define the residues involved in duck hepatitis B virus (DHBV) neutralization at the amino acid level, we have used a procedure combining monoclonal antibodies (MAbs) and overlapping octapeptides (Pepscan). Two neutralizing MAbs (SD20 and 900), specific for the pre-S protein were shown to reduce DHBV infectivity in vivo by 75 and 90%, respectively, while complete protection of ducklings was achieved with a polyclonal antiserum raised against the bacterially expressed first 131 amino acids of the DHBV pre-S region (DHBpre-S). Using fusion polypeptides, the binding sites of these MAbs were localized between aa 77 and 100 on pre-S protein. We have used octapeptides spanning the pre-S sequence from aa 64 to 115 for fine mapping of these epitopes. Within the sequence scanned, the polyclonal anti-DHBpre-S antiserum recognized a region exclusively limited to the residues E82-K95, suggesting immunodominance of this region in the sequence aa 64-115. The epitope recognized by Mab 900 was mapped within the same region, whereas the epitope recognized by Mab SD20 was localized downstream from this region. To define the amino acids essential for binding to the highly neutralizing Mab 900, we have used single amino acid replacement and demonstrated that two residues Q87 and W88 were important for antibody recognition.
Chassot
S
S
INSERM U 271, Lyon, France.
Lambert
V
V
Kay
A
A
Godinot
C
C
Roux
B
B
Trepo
C
C
Cova
L
L
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000914:Antibodies, Viral / Q000276:immunology; D000956:Antigens, Viral / Q000276:immunology*; D004372:Ducks; D000939:Epitopes; D004371:Hepatitis Virus, Duck / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D009842:Oligopeptides / Q000276:immunology; D017433:Protein Structure, Secondary; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000276:immunology*; D014764:Viral Proteins / Q000737:chemistry / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral; D000939:Epitopes; D009842:Oligopeptides; C077433:Pre-S protein, Duck hepatitis B virus; D014759:Viral Envelope Proteins; D014764:Viral Proteins