A Nucleophilic Activity-Based Probe Enables Profiling of PLP-Dependent Enzymes

Scott I Brody, Joseph A. Buonomo, Moyosore O Orimoloye, Ziyi Jia, Sachin Sharma, Christopher D. Brown, Anthony D. Baughn, Courtney C. Aldrich

Research output: Contribution to journalArticlepeer-review


PLP-dependent enzymes represent an important class of highly “druggable” enzymes that perform a wide array of critical reactions to support all organisms. Inhibition of individual members of this family of enzymes has been validated as a therapeutic target for pathologies ranging from infection with Mycobacterium tuberculosis to epilepsy. Given the broad nature of the activities within this family of enzymes, we envisioned a universally acting probe to characterize existing and putative members of the family that also includes the necessary chemical moieties to enable activity-based protein profiling experiments. Hence, we developed a probe that contains an N-hydroxyalanine warhead that acts as a covalent inhibitor of PLP-dependent enzymes, a linear diazirine for UV crosslinking, and an alkyne moiety to enable enrichment of crosslinked proteins. Our molecule was used to study PLP-dependent enzymes in vitro as well as look at whole-cell lysates of M. tuberculosis and assess inhibitory activity. The probe was able to enrich and identify LysA, a PLP-dependent enzyme crucial for lysine biosynthesis, through mass spectrometry. Overall, our study shows the utility of this trifunctional first-generation probe. We anticipate further optimization of probes for PLP-dependent enzymes will enable the characterization of rationally designed covalent inhibitors of PLP-dependent enzymes, which will expedite the preclinical characterization of these important therapeutic targets.

Original languageEnglish (US)
Article numbere202200669
Issue number7
StatePublished - Apr 3 2023

Bibliographical note

Publisher Copyright:
© 2023 The Authors. ChemBioChem published by Wiley-VCH GmbH.


  • PLP-dependent enzymes
  • activity-based protein profilin
  • chemical genetics
  • chemoproteomics
  • probes

PubMed: MeSH publication types

  • Journal Article


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