Mass Spectrometry-Based Top-Down Proteomics in Nanomedicine: Proteoform-Specific Measurement of Protein Corona

Conventional mass
spectrometry (MS)-based bottom-up proteomics
(BUP) analysis of the protein corona [i.e., an evolving layer of biomolecules,
mostly proteins, formed on the surface of nanoparticles (NPs) during
their interactions with biomolecular fluids] enabled the nanomedicine
community to partly identify the biological identity of NPs. Such
an approach, however, fails to pinpoint the specific proteoformsdistinct
molecular variants of proteins in the protein corona. The proteoform-level
information could potentially advance the prediction of the biological
fate and pharmacokinetics of nanomedicines. Recognizing this limitation,
this study pioneers a robust and reproducible MS-based top-down proteomics
(TDP) technique for characterizing proteoforms in the protein corona.
Our TDP approach has successfully identified about 900 proteoforms
in the protein corona of polystyrene NPs, ranging from 2 to 70 kDa,
revealing proteoforms of 48 protein biomarkers with combinations of
post-translational modifications, signal peptide cleavages, and/or
truncationsdetails that BUP could not fully discern. This
advancement in MS-based TDP offers a more advanced approach to characterize
NP protein coronas, deepening our understanding of NPs’ biological
identities. We, therefore, propose using both TDP and BUP strategies
to obtain more comprehensive information about the protein corona,
which, in turn, can further enhance the diagnostic and therapeutic
efficacy of nanomedicine technologies.