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Updated: Jan 10, 2026

Millisecond Hydrogen/Deuterium-Exchange Mass Spectrometry for the Study of Alpha-Synuclein Structural Dynamics Under Physiological Conditions
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HXMS: a standardized file format for HX/MS data.

Kyle C Weber1, Chenlin Lu1, Roberto Vera Alvarez2

  • 1Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA.

Biorxiv : the Preprint Server for Biology
|November 24, 2025
PubMed
Summary
This summary is machine-generated.

A new HXMS file format and PFLink software standardize hydrogen-deuterium exchange/mass spectrometry (HX/MS) data. This enables more quantitative analysis, better data sharing, and future machine learning applications for protein conformational studies.

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Area of Science:

  • Biochemistry and Structural Biology
  • Computational Biology and Bioinformatics

Background:

  • Hydrogen-deuterium exchange/mass spectrometry (HX/MS) is crucial for studying protein conformational dynamics.
  • Current HX/MS data formats lack standardization, hindering data analysis and information content, often using only centroid-level data instead of full isotopic mass spectra.

Purpose of the Study:

  • To develop a unified, standardized data format for HX/MS data.
  • To enhance the quantitative analysis and data sharing capabilities within the HX/MS community.

Main Methods:

  • Introduction of the HXMS file format, designed to be lightweight, scalable, and human-readable.
  • Development of PFLink, a Python package for converting existing HX/MS data files to the HXMS format.
  • Inclusion of support for isotopic mass envelopes, experimental time-courses, PTMs, and replicates.

Main Results:

  • The HXMS format preserves comprehensive isotopic and temporal data, unlike centroid-level formats.
  • PFLink facilitates seamless conversion of data from common HX/MS software to the standardized HXMS format.
  • The new format and tool support advanced analyses, including multimodal distributions and experimental replicates.

Conclusions:

  • The HXMS format and PFLink package offer a standardized solution for HX/MS data representation and analysis.
  • These developments will improve data sharing, quantitative accuracy, and enable future machine learning applications in protein science.
  • PFLink is publicly available, promoting wider adoption and collaboration in the HX/MS field.