Internal Fragments Enhance Middle-Down Mass Spectrometry Structural Characterization of Monoclonal Antibodies and Antibody-Drug Conjugates.

ABSTRACT: Monoclonal antibodies (mAbs) and antibody-drug conjugates (ADCs) are important large biotherapeutics (∼150 kDa) and high structural complexity that require extensive sequence and structure characterization. Middle-down mass spectrometry (MD-MS) is an emerging technique that sequences and maps subunits larger than those released by trypsinolysis. It avoids potentially introducing artifactual modifications that may occur in bottom-up MS while achieving higher sequence coverage compared to top-down MS. However, returning complete sequence information by MD-MS is still challenging. Here, we show that assigning internal fragments in direct infusion MD-MS of a mAb and an ADC substantially improves their structural characterization. For MD-MS of the reduced NIST mAb, including internal fragments

SUBMITTER: Wei B 

PROVIDER: S-EPMC11001303 | biostudies-literature | 2024 Feb

REPOSITORIES: biostudies-literature