PXD077479-1

PXD077479 is an original dataset announced via ProteomeXchange.

Title	Droplet microfluidic hydrogen/deuterium exchange for investigating protein dynamics with millisecond precision exchange
Description	Hydrogen/deuterium exchange (HDX) methods for studying protein dynamics would benefit from millisecond-scale incubations to probe intrinsically disordered proteins, highly dynamic regions and conformation changes. Here we investigate droplet microfluidics for rapid mixing to trigger D2O labelling, uniform incubations and rapid droplet merging for acid quenching in advance of mass spectrometry. A surfactant-free merging approach combining expansion elements for synchronised droplet collision proved robust. The high diffusive flux of D2O and protons enable microsecond mixing to trigger and arrest D2O labelling, respectively, affording the possibility of single millisecond incubations. Droplet HDX processors were used to measure the fast uptake characteristics of a model peptide. Forward exchange measurements demonstrate D2O labelling to be the rate-limiting step, in essence defining 10 milliseconds as the minimum practical incubation time for proteins in typical physiological conditions. With the ability to access millisecond time scales the fast dynamics of calmodulin, a model of calcium-triggered allostery with rapid conformational switching, was investigated. At 10 milliseconds, we could observe significant deuterium uptake within the well-defined EF-hand motifs (Ca2+ binding sites). These findings demonstrate millisecond HDX enabled by droplet microfluidics allows areas of heightened plasticity to be detected within a stably folded protein.
HostingRepository	PRIDE
AnnounceDate	2026-04-22
AnnouncementXML	Submission_2026-04-22_09:15:27.696.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Alistair Bailey
SpeciesList	scientific name: Bos taurus (Bovine); NCBI TaxID: NEWT:9913;
ModificationList	No PTMs are included in the dataset
Instrument	Xevo G2-XS QTof

Revision	Datetime	Status	ChangeLog Entry
0	2026-04-22 07:48:45	ID requested
⏵ 1	2026-04-22 09:15:28	announced

Dataset with its publication pending

submitter keyword: calmodulin,HDX-MS, droplet microfluidics

Eamonn Reading
contact affiliation	School of Biological Sciences, Building 85, University of Southampton, Southampton, SO17 1BJ
contact email	E.Reading@soton.ac.uk
lab head
Alistair Bailey
contact affiliation	University of Southampton
contact email	ab604@soton.ac.uk
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/04/PXD077479

PRIDE project URI

• PRIDE
  1. PXD077479
     1. Label: PRIDE project
     2. Name: Droplet microfluidic hydrogen/deuterium exchange for investigating protein dynamics with millisecond precision exchange