Journal Article

PUBDB-2023-06851

http://join2-wiki.gsi.de/foswiki/pub/Main/Artwork/join2_logo100x88.png Small conformational changes in IgG1 detected as acidic charge variants by cation exchange chromatography

Fukuda, M. (Corresponding author)Extern*EMBL* ; Graewert, M. A.EMBL* ; Jeffries, C. M.EMBL* ; Svergun, D. I.EMBL* ; Yamazaki, T. ; Koga, A. ; Yamanaka, Y.

2023
Elsevier San Diego, Calif.

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Please use a persistent id in citations: doi:10.1016/j.ab.2023.115302  doi:10.3204/PUBDB-2023-06851

Abstract: Fully characterizing the post-translational modifications present in charge variants of therapeutic monoclonal antibodies (mAbs), particularly acidic variants, is challenging and remains an open area of investigation. In this study, to test the possibility that chromatographically separated acidic fractions of therapeutic mAbs contain conformational variants, we undertook a mAb refolding approach using as a case study an IgG1 that contains many unidentified acidic peaks with few post-translational modifications, and examined whether different acidic peak fractions could be generated corresponding to these variants. The IgG1 drug substance was denatured by guanidine hydrochloride, without a reducing agent present, and gradually refolded by stepwise dialysis against arginine hydrochloride used as an aggregation suppressor. Each acidic chromatographic peak originally contained in the IgG1 drug substance was markedly increased by this stepwise refolding process, indicating that these acidic variants are conformational variants. However, no conformational changes were detected by small-angle X-ray scattering experiments for the whole IgG1, indicating that the conformational changes are minor. Chromatographic, thermal and fluorescence analyses suggested that the conformational changes are a localized denaturation effect centred around the aromatic amino acid regions. This study provides new insights into the characterization of acidic variants that are currently not fully understood.

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Contributing Institute(s):

1. EMBL (EMBL)
2. EMBL-User (EMBL-User)
3. DOOR-User (DOOR ; HAS-User)

Research Program(s):

1. 6G3 - PETRA III (DESY) (POF4-6G3) (POF4-6G3)

Experiment(s):

1. PETRA Beamline P12 (PETRA III)

Appears in the scientific report 2023

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Database coverage:
; ; BIOSIS Previews ; Clarivate Analytics Master Journal List ; Current Contents - Life Sciences ; Ebsco Academic Search ; IF < 5 ; JCR ; Nationallizenz ; SCOPUS ; Web of Science Core Collection

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