{"doi":"10.1021/jasms.3c00187","title":"Top-Down Protein Analysis by Tandem-Trapped Ion Mobility Spectrometry/Mass Spectrometry (Tandem-TIMS/MS) Coupled with Ultraviolet Photodissociation (UVPD) and Parallel Accumulation/Serial Fragmentation (PASEF) MS/MS Analysis","abstract":"\"Top-down\" proteomics analyzes intact proteins and identifies proteoforms by their intact mass as well as the observed fragmentation pattern in tandem mass spectrometry (MS/MS) experiments. Recently, hybrid ion mobility spectrometry-mass spectrometry (IM/MS) methods have gained traction for top-down experiments, either by allowing top-down analysis of individual isomers or alternatively by improving signal/noise and dynamic range for fragment ion assignment. We recently described the construction of a tandem-trapped ion mobility spectrometer/mass spectrometer (tandem-TIMS/MS) coupled with an ultraviolet (UV) laser and demonstrated a proof-of-principle for top-down analysis by UV photodissociation (UVPD) at 2-3 mbar. The present work builds on this with an exploration of a top-down method that couples tandem-TIMS/MS with UVPD and parallel-accumulation serial fragmentation (PASEF) MS/MS analysis. We first survey types and structures of UVPD-specific fragment ions generated in the 2-3 mbar pressure regime of our instrument. Notably, we observe UVPD-induced fragment ions with multiple conformations that differ from those produced in the absence of UV irradiation. Subsequently, we discuss how MS/MS spectra of top-down fragment ions lend themselves ideally for probability-based scoring methods developed in the bottom-up proteomics field and how the ability to record automated PASEF-MS/MS spectra resolves ambiguities in the assignment of top-down fragment ions. Finally, we describe the coupling of tandem-TIMS/MS workflows with UVPD and PASEF-MS/MS analysis for native top-down protein analysis.","journal":"Journal of the American Society for Mass Spectrometry","year":2023,"id":329490,"datarank":0.4493598410330987,"base_score":2.995732273553991,"endowment":2.995732273553991,"self_citation_contribution":0.4493598410330987,"citation_network_contribution":0.0,"self_endowment_contribution":0.4493598410330987,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":19,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9595,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2023-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":566150,"name":"Mark E. Ridgeway","orcid":"0000-0002-4700-2535","position":1,"is_corresponding":false},{"id":671040,"name":"Christopher A. Wootton","orcid":"0000-0002-3647-2611","position":2,"is_corresponding":false},{"id":671039,"name":"Alina Theisen","orcid":"0000-0002-0216-8582","position":3,"is_corresponding":false},{"id":353220,"name":"Erin Panczyk","orcid":"0000-0003-3779-6738","position":4,"is_corresponding":false},{"id":890816,"name":"Florian Meier","orcid":"0000-0003-4729-175X","position":5,"is_corresponding":false},{"id":89215,"name":"Melvin A. Park","orcid":"0009-0004-1448-2896","position":6,"is_corresponding":false},{"id":671041,"name":"Christian Bleiholder","orcid":"0000-0002-4211-1388","position":7,"is_corresponding":false},{"id":671035,"name":"Fanny C. Liu","orcid":"0000-0003-1403-7114","position":0,"is_corresponding":true}],"reference_count":93,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-19T01:09:01.509601Z","pmid":"37638640","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}