PXD061452-1

PXD061452 is an original dataset announced via ProteomeXchange.

Title	Self-packed size-exclusion columns enable versatile high-throughput native, top-down, and ion-mobility-mass spectrometry studies on proteins and complexes.
Description	Native MS (nMS) is a key structural biology technique that makes it possible to study intact proteins and their interactions. Unfortunately, non-volatile salts are incompatiblewith nMS, which demands a laborious desalting procedure. Non-denaturing size exclusion chromatography (SEC) allows both rapid desalting and separation and has previously been explored for nMS automation. However, SEC at conventional scale requires rather large sample amounts as well as harsh ESI conditions, which can cause protein unfolding. Capillary LC allows softer conditions; however, the few commercially available SEC columns appropriate for this flow rate are prohibitively expensive for many laboratories. Existing protocols for packing buffer exchange columns rely on specialized equipment and/or result in columns that have limited capacity for size-based protein separation. Here, we present self-packed miniaturized SEC columns with different stationary phases and customizable dimensions. The columns, produced via slurry packing with an ordinary LC pump were used across a range of samples in several applications including nMS, top-down MS (TDMS), ligand screening, and ion mobility (IM)-MS. Native separation allowed acquisition of data from samples containing more than one protein. We acquired native TDMS data of 3 proteins in 12 minutes, with up to 47% sequence coverage. IM-MS of alpha-synuclein at different charge states was measured in ca. 60 minutes (including calibrants), with results that match the literature. Finally, we used SEC-nMS to rapidly screen proteolysis-targeting chimera candidates and performed collision-induced unfolding (CIU) of a PROTAC-induced ternary complex. Through this work, we highlight the potential of SEC to support developments in structural MS.
HostingRepository	PRIDE
AnnounceDate	2025-06-29
AnnouncementXML	Submission_2025-06-29_10:29:31.970.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Edvaldo Maciiel
SpeciesList	scientific name: Escherichia coli; NCBI TaxID: 562; scientific name: Candida albicans (Yeast); NCBI TaxID: 5476; scientific name: Bos taurus (Bovine); NCBI TaxID: 9913; scientific name: Gallus gallus (Chicken); NCBI TaxID: 9031; scientific name: Homo sapiens (Human); NCBI TaxID: 9606; scientific name: Equus caballus (Horse); NCBI TaxID: 9796;
ModificationList	No PTMs are included in the dataset
Instrument	Synapt MS; nanoACQUITY UPLC

Revision	Datetime	Status	ChangeLog Entry
0	2025-03-04 05:48:14	ID requested
⏵ 1	2025-06-29 10:29:32	announced

Maciel EVS, Habeck T, Meyners C, Lermyte F, Self-packed size-exclusion columns enable versatile high-throughput native, top-down, and ion mobility-mass spectrometry studies on proteins and complexes. Talanta, 291():127868(2025) [pubmed]

10.1016/j.talanta.2025.127868;

submitter keyword: ligand screening, top-down proteomics, native mass spectrometry,Size-exclusion chromatography, PROTAC, ion mobility

Frederik Lermyte
contact affiliation	Clemens Schöpf Institute, Department of Chemistry, Technical University of Darmstadt, Peter-Grünberg-Straße 4, 64287 Darmstadt, Germany
contact email	frederik.lermyte@tu-darmstadt.de
lab head
Edvaldo Maciiel
contact affiliation	Technical University of Darmstadt
contact email	edvaldovasconcelossm@gmail.com
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD061452
     1. Label: PRIDE project
     2. Name: Self-packed size-exclusion columns enable versatile high-throughput native, top-down, and ion-mobility-mass spectrometry studies on proteins and complexes.