PXD051986-1

PXD051986 is an original dataset announced via ProteomeXchange.

Title	Screening privileged alkylguanidinium motifs under host-mimicking conditions reveals a novel antibiotic with an unconventional mode of action
Description	Screening large molecule libraries against pathogenic bacteria is often challenged by a low hit rate due to limited uptake, underrepresentation of antibiotic structural motifs and assays which do not resemble the infection conditions. To address these limitations, we here screen a library focused on alkylguanidinium compounds, a structural motif associated with antibiotic activity and enhanced uptake, under host-mimicking infection conditions against a panel of disease-associated bacteria. Several hit molecules were obtained with activities against Gram-positive as well as Gram-negative bacteria highlighting the fidelity of the general concept. Based on the best antibiotic activity, we selected one compound (L15) for in-depth mode of action studies. L15 exhibited bactericidal activity against Staphylococcus aureus USA300 Lac (JE2) with a minimum inhibitory concentration of 1.5 µM whereby a structure-activity relationship study with 18 derivatives proved the necessity of the guanidinium motif for antibiotic activity. Electron microscopy studies and depolarization assays at high L15 concentrations showed an impact on the cell membrane and integrity which was absent at the MIC concentration. Sequencing of L15-resistant strains revealed a mutation in an efflux pump but did not provide hints for direct targets. We thus performed affinity-based protein profiling with a L15 probe and identified signal peptidase IB (SpsB) as the most promising hit. Validation by activity assays, binding site identification and molecular docking demonstrated SpsB overactivation by L15, similar to a previously reported activator molecule that dysregulates protein secretion of this essential enzyme. Overall, this study highlights the need for unconventional screening strategies to identify novel antibiotics.
HostingRepository	PRIDE
AnnounceDate	2024-07-17
AnnouncementXML	Submission_2024-07-17_12:38:08.595.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Nina Bach
SpeciesList	scientific name: Staphylococcus aureus; NCBI TaxID: 1280;
ModificationList	iodoacetamide derivatized residue
Instrument	Orbitrap Eclipse; timsTOF Pro

Revision	Datetime	Status	ChangeLog Entry
0	2024-05-04 15:10:59	ID requested
⏵ 1	2024-07-17 12:38:08	announced

10.1021/JACSAU.4C00449;

submitter keyword: antibiotics, chemical proteomics, SAR, MoA studies,Guanidinium

Stephan Axel Sieber
contact affiliation	TUM School of Natural Sciences, Department Biosciences, Chair of Organic Chemistry II, Center for Functional Protein Assemblies (CPA), Technical University Munich (TUM), Ernst-Otto-Fischer Str. 8, Garching, 85748, Germany
contact email	stephan.sieber@tum.de
lab head
Nina Bach
contact affiliation	TU München
contact email	nina.bach@tum.de
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD051986
     1. Label: PRIDE project
     2. Name: Screening privileged alkylguanidinium motifs under host-mimicking conditions reveals a novel antibiotic with an unconventional mode of action