An industry-focused top-down mass spectrometry (TDMS) platform for resolving common ambiguities in protein characterization has been introduced. Bailey et al., from the Russell lab, developed a flow-programmed denaturing online buffer exchange (fp)dOBE method, providing extended MS/MS acquisition time with improved micro-flow ESI sensitivity. They also implemented an innovative data analysis technique using composite proteoform spectral match (cPrSM) results, accumulating fragment data from different scan types across multiple injections.

The study used α-lactalbumin as a model protein to demonstrate the platform’s capabilities. The researchers compared data-dependent acquisition (DDA) and targeted TDMS approaches, evaluating the impact of MS1 quality and fragmentation parameters. They showed that targeted methods could significantly increase the number of MS2 microscans acquired per proteoform.

This study demonstrates a streamlined, high-quality TDMS platform that can enhance protein characterization workflows in industrial settings.

An IonOpticks Aurora Ultimate 25×75 C18 UHPLC column was used for peptide mapping, showcasing its effectiveness in supporting comprehensive protein analysis.

Publication
Proteomics

Authors

Aaron O. Bailey, Kenneth R. Durbin, Matthew T. Robey, Lee K. Palmer, William K. Russell

Title

Filling the gaps in peptide maps with a platform assay for top‐down characterization of purified protein samples