Work

In‐Source Collision‐Induced Dissociation (CID) Improves Higher‐Energy Collisional Dissociation (HCD)‐Dependent Fragmentation of ADP‐Ribosyl Peptides

Year: 2024

Type: article

Abstract: ABSTRACT Rationale ADP‐ribosylation is a posttranslational modification whose higher‐energy collisional dissociation (HCD) products are dominated by complete or partial modification losses, complicating peptide sequencing and acceptor site localization efforts. We tested whether in‐source collision‐induced dissociation (CID) performed on a quadrupole‐Orbitrap could convert ADPr to the smaller phosphoribose‐H 2 O derivative to facilitate HCD‐dependent peptide sequencing. Methods ADP‐ribosyl (ADPr) peptides derived from the human macrophage‐like cell line THP‐1 were analyzed on a quadrupole‐Orbitrap. We monitored the dissociation of ADPr (+ 541.061 Da) to phosphoribosyl‐H 2 O (+ 193.997 Da) peptides while varying the source and high‐field asymmetric waveform ion mobility mass spectrometry (FAIMS) compensation voltages. Xcorr and ptmRS were used to evaluate peptide sequencing and acceptor site confidence, respectively. Phosphoribosyl‐H 2 O acceptor sites were compared with those determined by electron‐transfer higher‐energy collision dissociation (EThcD), performed on a quadrupole‐ion trap‐Orbitrap. Results In‐source CID of ADPr peptides to their phosphoribosyl‐H 2 O derivatives increased with increasing source voltage (up to 50 V), as judged by monitoring the corresponding modification loss ([adenosine monophosphate/AMP] + ) and the number of identified phosphoribosyl‐H 2 O peptide identifications. The average Xcorr increased from 1.36 (ADPr) to 2.26 (phosphoribosyl‐H 2 O), similar to that achieved with EThcD for ADPr peptides (2.29). The number of high‐confidence acceptor sites (> 95%) also increased, from 31% (ADPr) to 70% (phosphoribosyl‐H 2 O), which was comparable to EThcD (70%). Conclusions In‐source CID converts ADP‐ribosyl to phosphoribosyl‐H 2 O peptides that are more amenable to HCD‐dependent peptide sequencing, providing an alternative method for acceptor site determination when ETD‐based methods are not available.

Source: Rapid Communications in Mass Spectrometry

DOI: 10.1002/rcm.9961

Authors: Taku Kasai, Yuto Nakamura, Masanori Aikawa, Sasha A. Singh

Institutions: Brigham and Women's Hospital, Harvard University

Language: English

Topic: Mass Spectrometry Techniques and Applications

Subfield: Spectroscopy

Field: Chemistry

Domain: Physical Sciences

Open Access status: hybrid

Estimated APC paid: $4,840

Funders: Kowa Company, National Institutes of Health, National Heart, Lung, and Blood Institute