Improvement of neutralizing activity of human scFv antibodies against hepatitis B virus binding using CDR3 V(H) mutant library.
CDR3 of the heavy-chain variable region of immunoglobulin is a region in which somatic mutation occurs heavily after secondary antibody response, resulting in an affinity maturation of antibodies in vivo. The aim of this study was to improve the affinity of a human single-chain variable fragment (scFv) specific for pre-S1 of hepatitis B virus (HBV) by introducing random mutagenesis in CDR3 variable region of heavy chain (V(H)) of the parental scFv clone 1E4. By using a BIAcore for panning and screening, we have selected three clones (A9, B2, and B9) with lower highest affinity (K(D)) than 1E4. Affinities of selected clones ranged from 1.7 x 10(7) mol/L to 6.3 x 10(8) mol/L, which were increased by factors of 1.4 to 4.0, respectively, compared to the parental clone. Binding inhibition assay using flow cytometry and polymerase chain reaction revealed that B2 (6.4 x 10(8) mol/L) had a higher neutralizing activity against pre-S1 or HBV virion binding to liver cell line. This anti-pre-S1 scFv can be considered as a potential therapeutic tool for a passive immunotherapy for HBV infection.
Department of Microbiology, College of Medicine and Center for Viral Disease Research, Inje University, Pusan, Republic of Korea.
Sae-Gwang
SG
Park
Young-Joo
YJ
Jung
Young-Yi
YY
Lee
Cheul-Min
CM
Yang
Ik-Jung
IJ
Kim
Jun-Ho
JH
Chung
Ik-Sang
IS
Kim
Youn-Jae
YJ
Lee
Sung-Jae
SJ
Park
Jeong-Nyeo
JN
Lee
Su-Kil
SK
Seo
Yong-Hong
YH
Park
In-Hak
IH
Choi
D000595:Amino Acid Sequence; D006510:Hepatitis B Antibodies / Q000276:immunology*; D006515:Hepatitis B virus / Q000276:immunology* / Q000378:metabolism*; D006801:Humans; D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D009500:Neutralization Tests; D019151:Peptide Library; D011485:Protein Binding
D006510:Hepatitis B Antibodies; D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region; D019151:Peptide Library