Dataset Information

Epitope and paratope mapping of a SUMO-remnant antibody using cross-linking mass spectrometry and molecular docking

ABSTRACT: The small ubiquitin-like modifier (SUMO) is an important post-translational modification that regulates the function of various proteins essential for DNA damage repair, genome integrity and cell homeostasis. To identify protein SUMOylation effectively, an enrichment step is necessary, often requires exogenous gene expression in cells and immunoaffinity purification of SUMO-remnant peptides following tryptic digestion. Previously, an antibody was developed to enrich tryptic peptides containing the remnant NQTGG on the receptor lysine, though the specifics of the structural interaction motif remained unclear. Here, we conducted de novo sequencing by mass spectrometry to analyze the SUMO-remnant antibody and performed paratope mapping using cross-linking experiments to identify key interacting residues within the complementarity-determining regions (CDRs). Additional cross-linking experiments were performed using SUMOylated peptides and high-field asymmetric waveform ion mobility spectrometry (FAIMS) to enhance sensitivity and confirm interactions at the paratope interface. Our comprehensive analyses have provided a detailed understanding of the structural interaction motifs of the SUMO-remnant antibody, offering valuable insights into the antibody’s specificity and binding mechanisms. This enhanced understanding paves the way for improved methodologies in studying protein SUMOylation and its regulatory roles in cellular processes.

INSTRUMENT(S): Orbitrap Fusion, Orbitrap Exploris 480, Q Exactive HF

ORGANISM(S): Oryctolagus Sp. 'rabbit_od'

SUBMITTER: Simon Comtois-Marotte

LAB HEAD: Pierre Thibault

PROVIDER: PXD055085 | Pride | 2025-02-19

REPOSITORIES: pride

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Dataset's files

Source:

			Action	DRS
	DSS_antiKNQTGG_nonspeci.zip	Other
	Sumo_Ab_1ug_AspN.raw	Raw
	Sumo_Ab_1ug_AspN.raw.mzxml	Mzxml
	Sumo_Ab_1ug_Chymotrypsin.raw	Raw
	Sumo_Ab_1ug_Chymotrypsin.raw.mzxml	Mzxml

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Publications

Epitope and Paratope Mapping of a SUMO-Remnant Antibody Using Cross-Linking Mass Spectrometry and Molecular Docking.

Comtois-Marotte Simon S Bonneil Éric É Li Chongyang C Smith Matthew J MJ Thibault Pierre P

Journal of proteome research 20250218

The small ubiquitin-like modifier (SUMO) is an important post-translational modification that regulates the function of various proteins essential for DNA damage repair, genome integrity, and cell homeostasis. To identify protein SUMOylation effectively, an enrichment step is necessary, often requiring exogenous gene expression in cells and immunoaffinity purification of SUMO-remnant peptides following tryptic digestion. Previously, an antibody was developed to enrich tryptic peptides containing ...[more]

PMID: 39965925

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