193 nm ultraviolet photodissociation for the characterization of singly charged proteoforms generated by MALDI

14 December 2022, Version 1
This content is a preprint and has not undergone peer review at the time of posting.

Abstract

MALDI imaging allows for the near-cellular profiling of proteoforms directly from microbial colonies, or plant and mammalian tissues. Despite detecting hundreds of proteoforms, identification of unknowns with only intact mass information remains a distinct challenge, even at with high resolution and mass accuracy. To this end many supplementary methods have been used to create experimental databases for accurate mass matching, including bulk or spatially resolved bottom-up and/or top-down proteomics. Herein we describe the application of 193 nm ultraviolet photodissociation (UVPD) for fragmentation of quadrupole isolated singly charged ubiquitin (m/z 8565) by MALDI-UVPD on an UHMR HF Orbitrap. This platform permitted the high-resolution accurate mass measurement of not just terminal fragments, but also large internal fragments. The outlined workflow demonstrated the feasibility of high resolution top-down analysis of MALDI generated protein ions and the potential towards more comprehensive characterization of proteoforms in MALDI imaging applications.

Keywords

ultraviolet photodissociation
UVPD
matrix-assisted laser desorption/ionization
MALDI
dissociation
top-down proteomics
ubiquitin

Supplementary materials

Title
Description
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Title
Supplementary Information
Description
Includes extended discussion about MALDI-UVPD implementation, tuning, and use with further analyses of attenuation of excimer pulse energies through optics, and several different representations of the annotated fragmentation data of various levels of microscan averaging.
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Supplementary Data
Description
A zipped folder containing .RAW files corresponding to the total of 1500 microscan averages as described within this note. From an isolation acquisition with no fluence, to an experiment with 5.5 mJ pulse energy.
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