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Related Experiment Video

Updated: Jan 14, 2026

Analyzing Protein Dynamics Using Hydrogen Exchange Mass Spectrometry

Analyzing Protein Dynamics Using Hydrogen Exchange Mass Spectrometry

Published on: November 29, 2013

Site-resolved energetic information from HX-MS experiments.

Chenlin Lu¹, Malcolm L Wells¹, Andrew Reckers¹

¹Department of Biochemistry and Molecular Biophysics, Columbia University Irving Medical Center, New York, NY, USA.

Nature Chemical Biology

|October 17, 2025

Summary

This summary is machine-generated.

PIGEON-FEATHER analyzes protein dynamics from HX-MS data, revealing distinct enzyme ensembles and inhibitor binding. This method solves longstanding questions in protein evolution and function.

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Last Updated: Jan 14, 2026

Analyzing Protein Dynamics Using Hydrogen Exchange Mass Spectrometry

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Published on: November 29, 2013

A Hydrogen-Deuterium Exchange Mass Spectrometry HDX-MS Platform for Investigating Peptide Biosynthetic Enzymes

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Time-resolved ElectroSpray Ionization Hydrogen-deuterium Exchange Mass Spectrometry for Studying Protein Structure and Dynamics

Time-resolved ElectroSpray Ionization Hydrogen-deuterium Exchange Mass Spectrometry for Studying Protein Structure and Dynamics

Published on: April 17, 2017

Area of Science:

Biophysics
Structural Biology
Computational Biology

Background:

Understanding protein conformational ensembles is crucial for function but difficult to achieve at high resolution.
Existing methods lack the resolution to capture subtle energetic differences in protein dynamics.

Purpose of the Study:

To introduce PIGEON-FEATHER, a novel computational method for high-resolution analysis of protein conformational ensembles.
To calculate ensemble free energies of opening (∆G_op) at single-amino-acid resolution using hydrogen exchange-mass spectrometry (HX-MS) data.

Main Methods:

Developed PIGEON-FEATHER, a Bayesian Monte Carlo sampling approach.
Applied the method to reconstruct HX-MS isotopic mass envelopes for proteins of various sizes.
Utilized HX-MS data to analyze protein dynamics and ensemble energetics.

Main Results:

Revealed distinct evolutionary pathways and conformational ensembles for Escherichia coli and human dihydrofolate reductases (ecDHFR and hDHFR).
Explained differential inhibitor binding to ecDHFR and hDHFR, resolving a key question in drug efficacy.
Mapped ligand-induced ensemble reweighting in the E. coli lac repressor, elucidating its transcriptional regulation mechanism.

Conclusions:

PIGEON-FEATHER provides high-resolution energetic insights into protein conformational ensembles from HX-MS data.
The method elucidates evolutionary divergence in protein function and differential drug responses.
PIGEON-FEATHER offers a powerful tool for understanding complex biological systems, including protein-DNA interactions and transcriptional regulation.