Open AccessCharacterization of Humanized Antibodies Secreted byAspergillus niger
Author(s) -
Michael P. Ward,
Cherry Lin,
Doreen Victoria,
Bryan P. Fox,
Judith A. Fox,
David L. Wong,
Hendrik J. Meerman,
Jeff P. Pucci,
Robin B. Fong,
Meng Heng,
Naoya Tsurushita,
Christine Gieswein,
Minha Park,
Huaming Wang
Publication year - 2004
Publication title -
applied and environmental microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.552
H-Index - 324
eISSN - 1070-6291
pISSN - 0099-2240
DOI - 10.1128/aem.70.5.2567-2576.2004
Subject(s) - antibody , aspergillus niger , fragment crystallizable region , biochemistry , glycosylation , avidity , mannose , chemistry , immunoglobulin light chain , microbiology and biotechnology , galactose , glycan , biology , glycoprotein , immunology
Two different humanized immunoglobulin G1(kappa) antibodies and an Fab' fragment were produced by Aspergillus niger. The antibodies were secreted into the culture supernatant. Both light and heavy chains were initially synthesized as fusion proteins with native glucoamylase. After antibody assembly, cleavage by A. niger KexB protease allowed the release of free antibody. Purification by hydrophobic charge induction chromatography proved effective at removing any antibody to which glucoamylase remained attached. Glycosylation at N297 in the Fc region of the heavy chain was observed, but this site was unoccupied on approximately 50% of the heavy chains. The glycan was of the high-mannose type, with some galactose present, and the size ranged from Hex(6)GlcNAc(2) to Hex(15)GlcNAc(2). An aglycosyl mutant form of antibody was also produced. No significant difference between the glycosylated antibody produced by Aspergillus and that produced by mammalian cell cultures was observed in tests for affinity, avidity, pharmacokinetics, or antibody-dependent cellular cytotoxicity function.