3 research outputs found
Global and Local Conformation of Human IgG Antibody Variants Rationalizes Loss of Thermodynamic Stability.
Immunoglobulin G (IgG) monoclonal antibodies (mAbs) are a major class of medicines, with high specificity and affinity towards targets spanning many disease areas. The antibody Fc (fragment crystallizable) region is a vital component of existing antibody therapeutics, as well as many next generation biologic medicines. Thermodynamic stability is a critical property for the development of stable and effective therapeutic proteins. Herein, a combination of ion-mobility mass spectrometry (IM-MS) and hydrogen/deuterium exchange mass spectrometry (HDX-MS) approaches have been used to inform on the global and local conformation and dynamics of engineered IgG Fc variants with reduced thermodynamic stability. The changes in conformation and dynamics have been correlated with their thermodynamic stability to better understand the destabilising effect of functional IgG Fc mutations and to inform engineering of future therapeutic proteins.This is the author accepted manuscript. The final version is available from Wiley via http://dx.doi.org/10.1002/anie.20150722
Rate of Asparagine Deamidation in a Monoclonal Antibody Correlating with Hydrogen Exchange Rate at Adjacent Downstream Residues
Antibodies are an important class
of drugs, comprising more than half of all new FDA approvals. Therapeutic
antibodies must be chemically stable both in storage and <i>in
vivo</i>, following administration to patients. Deamidation is
a major degradation pathway for all natural and therapeutic proteins
circulating in blood. Here, the linkage between deamidation propensity
and structural dynamics is investigated by examining two antibodies
with differing specificities. While both antibodies share a canonical
asparagine-glycine (NG) motif in a structural loop, this is prone to
deamidation in one of the antibodies but not the other. We found that
the hydrogen-exchange rate at the adjacent two amides, often the autocatalytic
nucleophiles in deamidation, correlated with the rate of degradation.
This previously unreported observation was confirmed upon mutation
to stabilize the deamidation lability via a generally applicable orthogonal
engineering strategy presented here. We anticipate that the structural
insight into chemical degradation in full-length monoclonal antibodies
and the high-resolution hydrogen-exchange methodology used will have
broad application across biochemical study and drug discovery and
development