Top-down ion mobility/mass spectrometry reveals enzyme specificity: Separation and sequencing of isomeric proteoforms

Published: 15 November 2023| Version 1 | DOI: 10.17632/gghrfnj4kr.1
Francis Berthias


IMS-MS/MS data associated with the under-review paper: 10.22541/au.168908634.44353344/v1 Abstract: "We assessed enzyme specificity using a top-down ion mobility spectrometry (IMS) and tandem mass spectrometry (MS/MS) workflow. We successfully applied trapped IMS (TIMS) to investigate site-specific N-ε-acetylation of lysine residues of full-length histone H4 catalyzed by histone lysine acetyltransferase KAT8. We demonstrate that KAT8 exhibits a preference for N-ε-actylation of residue K16, while also installing N-ε-acetyl groups on residues K5 and K8 as the first degree of acetylation. Achieving TIMS resolving power values of up to 300, we fully separated mono-acetylated regioisomers (H4K5ac, H4K8ac, and H4K16ac). Each of these regioisomers produce unique MS/MS fragment ions, enabling estimation of their individual mobility distributions and the exact localization of the N-ε-acetylation sites. This study highlights the potential of top-down TIMS-MS/MS for conducting enzymatic assays at the intact protein level and, more generally, for separation and identification of isomeric proteoforms and precise PTM localization." Data were recorded on a timsTOF Pro platform (Bruker). The directory 'rawData_BrukerFormat' contains all the raw data from the instrument. The directory 'Extracted_Data' contains all the extracted data used to plot the figures in the draft paper."


Steps to reproduce

Data associated to under review paper: 10.22541/au.168908634.44353344/v1 Raw data was extracted using DataAnalysis Software (Bruker). The list of extracted b-fragment ions are provided in SI (a copy of the table of extracted ion with ppm are copied in the rawdatafolder)


Biologisk Institut Syddansk Universitet


Mass Spectrometry, Proteomics