PXD029832

PXD029832 is an original dataset announced via ProteomeXchange.

Title	Tailored pyridoxal probes unravel novel cofactor-dependent targets and antibiotic hits in critical bacterial pathogens
Description	Unprecedented bacterial targets are urgently needed for the development of novel antibiotics to overcome the current resistance crisis. Challenges include the limited uptake of compounds as well as prioritizing proteins for their druggability and functional relevance. Especially, the wealth of uncharacterized proteins represents an untapped source for novel targets. However, tools to decipher their function are largely lacking. We here utilize the systematic mining of pyridoxal phosphate dependent enzymes (PLP DEs) in bacteria to focus on a target class, which is known to bind ligands, accesses PLP via active transport from the media and is involved in crucial metabolic processes. For this, we systematically exploited the chemical space of the pyridoxal (PL) scaffold and obtained eight PL probes bearing modifications at various ring positions. These probes were subsequently tested for phosphorylation by cognate kinases and labelling of PLP DEs in clinically relevant Gram-positive (Staphylococcus aureus) as well as Gram-negative (Escherichia coli and Pseudomonas aeruginosa) strains. Overall, the coverage of this diverse set of probes along with refined labelling conditions not only exceeded the performance of a previous probe generation, it also provided a detailed map of binding preferences of certain structural motifs. Although originally conducted in mutant cells devoid of PLP de novo biosynthesis, we here demonstrate efficient PLP DE labelling also in a wild type strain. Overall, the profiling revealed several putative PLP DEs with unknown function, of which we exemplarily characterized five via in-depth enzymatic assays. Finally, we screened a panel of putative PLP binders for antibiotic activity and unravelled the targets of hit molecules via competitive profiling with our probes. Here, an uncharacterized enzyme, essential for bacterial growth, was assigned as PLP dependent cysteine desulfurase and confirmed to be inhibited by the marketed drug phenelzine. Our approach provides a basis for deciphering novel PLP DEs as essential antibiotic targets along with corresponding ways to decipher small molecule inhibitors.
HostingRepository	PRIDE
AnnounceDate	2022-08-12
AnnouncementXML	Submission_2022-08-12_09:17:15.101.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Martin Pfanzelt
SpeciesList	scientific name: Escherichia coli; NCBI TaxID: 562; scientific name: Pseudomonas aeruginosa; NCBI TaxID: 287; scientific name: Staphylococcus aureus; NCBI TaxID: 1280;
ModificationList	No PTMs are included in the dataset
Instrument	Orbitrap Fusion

Revision	Datetime	Status	ChangeLog Entry
0	2021-11-20 08:46:52	ID requested
⏵ 1	2022-08-12 09:17:16	announced

Pfanzelt M, Maher TE, Absmeier RM, Schwarz M, Sieber SA, Tailored Pyridoxal Probes Unravel Novel Cofactor-Dependent Targets and Antibiotic Hits in Critical Bacterial Pathogens. Angew Chem Int Ed Engl, 61(24):e202117724(2022) [pubmed]

submitter keyword: Pyridoxal phosphate dependent enzymes, chemical proteomics, enzyme characterization, antibiotic compound screening

Stephan Axel Sieber
contact affiliation	Chair of Organic Chemistry II TU München Ernst-Otto-Fischer-Straße 8 D-85748 Garching Germany
contact email	stephan.sieber@tum.de
lab head
Martin Pfanzelt
contact affiliation	Technical University of Munich
contact email	martin.pfanzelt@tum.de
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD029832
     1. Label: PRIDE project
     2. Name: Tailored pyridoxal probes unravel novel cofactor-dependent targets and antibiotic hits in critical bacterial pathogens