首页|Mapping conformational changes on bispecific antigen-binding biotherapeutic by covalent labeling and mass spectrometry
Mapping conformational changes on bispecific antigen-binding biotherapeutic by covalent labeling and mass spectrometry
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国家科技期刊平台
NETL
NSTL
万方数据
Biotherapeutic's higher order structure(HOS)is a critical determinant of its functional properties and conformational relevance.Here,we evaluated two covalent labeling methods:diethylpyrocarbonate(DEPC)-labeling and fast photooxidation of proteins(FPOP),in conjunction with mass spectrometry(MS),to investigate structural modifications for the new class of immuno-oncological therapy known as bis-pecific antigen-binding biotherapeutics(BABB).The evaluated techniques unveiled subtle structural changes occurring at the amino acid residue level within the antigen-binding domain under both native and thermal stress conditions,which cannot be detected by conventional biophysical techniques,e.g.,near-ultraviolet circular dichroism(NUV-CD).The determined variations in labeling uptake under native and stress conditions,corroborated by binding assays,shed light on the binding effect,and highlighted the potential of covalent-labeling methods to effectively monitor conformational changes that ultimately influence the product quality.Our study provides a foundation for implementing the developed tech-niques in elucidating the inherent structural characteristics of novel therapeutics and their conforma-tional stability.
Covalent labeling/footprintingLiquid chromatography-mass spectrometryFast photooxidation of proteinsDiethylpyrocarbonateHigher order structure characterization
国家科技期刊平台
NSTL
万方数据
