title: Effects of salts from the Hofmeister series on the conformational stability, aggregation propensity, and local flexibility of an IgG1 monoclonal antibody.
abstract: This work examines the effect of three anions from the Hofmeister series (sulfate, chloride, and thiocyanate) on the conformational stability and aggregation rate of an IgG1 monoclonal antibody (mAb) and corresponding changes in the mAb's backbone flexibility (at pH 6 and 25 °C). Compared to a 0.1 M NaCl control, thiocyanate (0.5 M) decreased the melting temperatures (Tm) for three observed conformational transitions within the mAb by 6-9 °C, as measured by differential scanning calorimetry. Thiocyanate also accelerated the rate of monomer loss at 40 °C over 12 months, as monitored by size exclusion chromatography. Backbone flexibility, as measured via H/D exchange mass spectrometry, increased in two segments in the CH2 domain with more subtle changes across several additional regions. Chloride (0.5 M) caused slight increases in the Tm values, small changes in aggregation rate, and minimal yet consistent decreases in flexibility across various domains with larger effects noted within the VL, CH1, and CH3 domains. In contrast, 0.5 M sulfate increased Tm values, had small stabilizing influences on aggregate formation over time, yet substantially increased the flexibility of two specific regions in the CH1 and VL domains. While thiocyanate-induced conformational destabilization of the mAb correlated with increased local flexibility of specific regions in the CH2 domain (especially residues 241-251 in the heavy chain), the stabilizing anion sulfate did not affect these CH2 regions.
authors.affiliation: Department of Pharmaceutical Chemistry, Macromolecule and Vaccine Stabilization Center, University of Kansas , Lawrence, Kansas 66047, United States.
authors.forename: Ranajoy
authors.initials: R
authors.lastname: Majumdar
authors.forename: Prakash
authors.initials: P
authors.lastname: Manikwar
authors.forename: John M
authors.initials: JM
authors.lastname: Hickey
authors.forename: Hardeep S
authors.initials: HS
authors.lastname: Samra
authors.forename: Hasige A
authors.initials: HA
authors.lastname: Sathish
authors.forename: Steven M
authors.initials: SM
authors.lastname: Bishop
authors.forename: C Russell
authors.initials: CR
authors.lastname: Middaugh
authors.forename: David B
authors.initials: DB
authors.lastname: Volkin
authors.forename: David D
authors.initials: DD
authors.lastname: Weis
mesh_terms: D000838:Anions; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry*; D008958:Models, Molecular; D011487:Protein Conformation; D055503:Protein Multimerization; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D012492:Salts; D013816:Thermodynamics
chemical_list: D000838:Anions; D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D012492:Salts