Conformational Difference in Human IgG2 Disulfide
Isoforms Revealed by Hydrogen/Deuterium Exchange Mass Spectrometry
- Publication date
- Publisher
Abstract
Both recombinant and natural human
IgG2 antibodies have several
different disulfide bond isoforms, which possess different global
structures, thermal stabilities, and biological activities. A detailed
mapping of the structural difference among IgG2 disulfide isoforms,
however, has not been established. In this work, we employed hydrogen/deuterium
exchange mass spectrometry to study the conformation of three major
IgG2 disulfide isoforms known as IgG2-B, IgG2-A1, and IgG2-A2 in two
recombinant human IgG2 monoclonal antibodies. By comparing the protection
factors between amino acid residues in isoforms B and A1 (the classical
form), we successfully identified several local regions in which the
IgG2-B isoform showed more solvent protection than the IgG2-A1 isoform.
On the basis of three-dimensional structural models of IgG2, these
identified regions were located on the Fab domains, close to the hinge,
centered on the side where the two Fab arms faced each other in spatial
proximity. We speculated that in the more solvent-protected B isoform,
the two Fab arms were brought into contact by the nonclassical disulfide
bonds, resulting in a more compact global structure. Loss of Fab domain
flexibility in IgG2-B could limit its ability to access cell-surface
epitopes, leading to reduced antigen binding potency. The A2 isoform
was previously found to have disulfide linkages similar to those of
the classical A1 isoform, but with different biophysical behaviors.
Our data indicated that, compared to IgG2-A1, IgG2-A2 had less solvent
protection in some heavy-chain Fab regions close the hinge, suggesting
that the A2 isoform had more flexible Fab domains