Local dynamics and their alteration by excipients modulate the global conformational stability of an lgG1 monoclonal antibody.
A molecular understanding of excipient effects on the interrelationship(s) between dynamics and conformational stability of proteins, such as monoclonal antibodies (mAbs), can be important for their pharmaceutical development. The current study examines stabilizing and destabilizing effects of excipients on the conformational stability and local dynamics of distinct solvent-exposed regions within an IgG1 monoclonal antibody (mAb-B). The principles of site-selective photoselection upon red-edge excitation, accompanied by acrylamide quenching of tryptophan fluorescence were employed in this study. The initiation of mAb-B thermal unfolding occurs by structural alterations in the more solvent-exposed regions of the antibody, which subsequently leads to a cascade of structural alterations in its relatively more solvent-shielded regions. In addition, an increase in internal dynamics of solvent-shielded regions made mAb-B more susceptible to thermally induced structural perturbations resulting in its global destabilization. Sucrose and arginine exert their stabilizing and destabilizing effects by predominantly influencing the conformational stability of solvent-exposed regions in mAb-B. The complex molecular effects of sucrose and arginine on local dynamics of different regions in mAb-B and their correlation with the protein's conformational stability are described within the pretransition range, at the onset temperature (T(onset)) and at the thermal melting temperature (T(M)).
Department of Pharmaceutical Chemistry, Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, Kansas 66047, USA.
Santosh V
SV
Thakkar
Jae Hyun
JH
Kim
Hardeep S
HS
Samra
Hasige A
HA
Sathish
Steven M
SM
Bishop
Sangeeta B
SB
Joshi
David B
DB
Volkin
C Russell
CR
Middaugh
D000911:Antibodies, Monoclonal / Q000737:chemistry*; D001120:Arginine / Q000737:chemistry; D005079:Excipients / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry*; D011487:Protein Conformation; D055550:Protein Stability; D058767:Protein Unfolding; D013395:Sucrose / Q000737:chemistry; D013696:Temperature
D000911:Antibodies, Monoclonal; D005079:Excipients; D007074:Immunoglobulin G; D013395:Sucrose; D001120:Arginine