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Interchain Disulfide Bonding in Human IgG2 Antibodies Probed by Site-Directed Mutagenesis
- Source :
- Biochemistry. 48:3755-3766
- Publication Year :
- 2009
- Publisher :
- American Chemical Society (ACS), 2009.
-
Abstract
- Human IgG2 exists as a mixture of disulfide-linked structural isoforms that can show different activities. To probe the contribution of specific cysteine residues to the formation of structural isoforms, we characterized a series of Cys-->Ser mutant IgG2 recombinant monoclonal antibodies, focused on the first C(H)1 cysteine and the first two hinge cysteines. These residues participate in the formation of structural isoforms that have been noted by nonreduced capillary sodium dodecyl sulfate polyacrylamide gel electrophoresis, reversed-phase high-performance liquid chromatography, and cation exchange chromatography. We show that single Cys-->Ser mutants can greatly reduce heterogeneous disulfide bonding in human IgG2 and maintain in vitro activity. The data demonstrate the feasibility of applying site-directed mutagenesis to reduce disulfide bond heterogeneity in human IgG2 while preserving the activity of this therapeutically important class of human antibodies.
- Subjects :
- Spectrometry, Mass, Electrospray Ionization
medicine.drug_class
Ion chromatography
Monoclonal antibody
Peptide Mapping
Biochemistry
Immunoglobulin kappa-Chains
chemistry.chemical_compound
Serine
medicine
Humans
Cysteine
Disulfides
Sodium dodecyl sulfate
Site-directed mutagenesis
Polyacrylamide gel electrophoresis
Cysteine metabolism
Mutagenesis
Antibodies, Monoclonal
Peptide Fragments
Recombinant Proteins
Amino Acid Substitution
chemistry
Immunoglobulin G
Mutagenesis, Site-Directed
Electrophoresis, Polyacrylamide Gel
Subjects
Details
- ISSN :
- 15204995 and 00062960
- Volume :
- 48
- Database :
- OpenAIRE
- Journal :
- Biochemistry
- Accession number :
- edsair.doi.dedup.....f47a4e710ff16421f907836b77233570