Proteoform Thursday: Benqian Wei presents “Internal Fragments in Top-Down and Middle-Down Mass Spectrometry: From Fundamentals to Applications”

April 25, 2024 11:00 AM in Eastern Time (US and Canada)

Top-down and middle-down mass spectrometry (TD- and MD-MS) have become prominent analytical tools for protein characterization. However, obtaining complete sequence coverage by TD-/MD-MS can be challenging, particularly for proteins greater than 30 kDa including therapeutic monoclonal antibodies (mAbs) and antibody-drug-conjugates (ADCs). Assigning non-canonical internal fragments can help address this limitation. In this talk, I will start by discussing the fundamental formation mechanism of internal fragments which aids the development of sequencing algorithms. Next, I will describe how we applied internal fragment analysis in TD- and MD-MS to recover nearly 100% of the sequence and reveal important disulfide connectivity information of an intact mAb. In addition, drug conjugation sites can also be determined for a heterogeneous lysine-linked ADC using this novel approach