PXD058094

PXD058094 is an original dataset announced via ProteomeXchange.

Title	Novel calcium-binding sites in the acylated segment control the folding and membrane penetration of RTX toxins
Description	The Repeats in ToXin (RTX) domain of RTX cytolysins harbors numerous hexacoordinated Ca2+-binding sites that facilitate cooperative and vectorial folding of the RTX β-rolls. Recent study identified Ca2+-binding sites in the acylated N-terminal extension of the RTX domain of Bordetella pertussis adenylate cyclase toxin (CyaA), where the side chains of residues D880, D918 and N936 coordinate two calcium ions. Substitution of N936 with a bulky leucine did not affect the essential acylation of internal lysines or the capacity of the CyaA N936L to bind the receptor CR3. However, this substitution impaired the calcium-dependent folding of the acylated segment and abolished the capacity of the toxin to penetrate cell membrane. Similarly, substitution of the homologous D639 residue strongly reduced the cytotoxicity of Escherichia coli α-hemolysin (HlyA). While the substitution of D918 with leucine had no effect on CyaA activity, the D880L variant enhanced cell binding, adenylate cyclase (AC) domain translocation to cell cytosol and both the specific hemolytic activity of the CyaA D880L on erythrocytes and the membrane activity in artificial lipid bilayers. Hydrogen/deuterium exchange revealed that the D880L substitution altered the folding of the acylated segment and reduced the deuteration accessibility of surafce residues in the two loops bearing the acyl chains. Additionally, substitution of the β-turn-forming glycine G934 in CyaA and G637 in HlyA impaired the folding of the acylated segment and decreased the cytotoxicity of both toxins. These results highlight the critical role of the acylated segment in the membrane penetration capacity of RTX toxins from Gram-negative pathogens.
HostingRepository	PRIDE
AnnounceDate	2026-04-22
AnnouncementXML	Submission_2026-04-22_06:56:16.978.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Zuzana Kalaninova
SpeciesList	scientific name: Bordetella pertussis; NCBI TaxID: NEWT:520;
ModificationList	N6-acylated L-lysine
Instrument	timsTOF Pro

Revision	Datetime	Status	ChangeLog Entry
0	2024-11-20 07:51:12	ID requested
⏵ 1	2026-04-22 06:56:17	announced

Dataset with its publication pending

ProteomeXchange project tag: Hydrogen Deuterium Exchange (HDX-MS)

submitter keyword: membrane penetration, H/D exchange,RTX toxin, folding, calcium binding site, acylated segment

Petr Man
contact affiliation	Institute of Microbiology - BioCeV, Academy of Sciences of the Czech Republic
contact email	pman@biomed.cas.cz
lab head
Zuzana Kalaninova
contact affiliation	Faculty of Science, Charles University
contact email	kalaninz@natur.cuni.cz
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/PXD058094

PRIDE project URI

• PRIDE
  1. PXD058094
     1. Label: PRIDE project
     2. Name: Novel calcium-binding sites in the acylated segment control the folding and membrane penetration of RTX toxins