Comparison of Bioorthogonal β-Lactone Activity-Based Probes for Selective Labeling of Penicillin-Binding Proteins

Nathaniel W. Brown, Joshua D. Shirley, Andrew P. Marshall, Erin E. Carlson

Research output: Contribution to journalArticlepeer-review

Abstract

Penicillin-binding proteins (PBPs) are a family of bacterial enzymes that are key components of cell-wall biosynthesis and the target of β-lactam antibiotics. Most microbial pathogens contain multiple structurally homologous PBP isoforms, making it difficult to target individual PBPs. To study the roles and regulation of specific PBP isoforms, a panel of bioorthogonal β-lactone probes was synthesized and compared. Fluorescent labeling confirmed selectivity, and PBPs were selectively enriched from Streptococcus pneumoniae lysates. Comparisons between fluorescent labeling of probes revealed that the accessibility of bioorthogonal reporter molecules to the bound probe in the native protein environment exerts a more significant effect on labeling intensity than the bioorthogonal reaction used, observations that are likely applicable beyond this class of probes or proteins. Selective, bioorthogonal activity-based probes for PBPs will facilitate the activity-based determination of the roles and regulation of specific PBP isoforms, a key gap in knowledge that has yet to be filled.

Original languageEnglish (US)
Pages (from-to)193-202
Number of pages10
JournalChemBioChem
Volume22
Issue number1
DOIs
StatePublished - Jan 5 2021

Bibliographical note

Funding Information:
The authors thank the M. Winkler Lab at Indiana University for providing strains, especially T. Tsui for extensive discussions, and the Carlson Lab for helpful discussion and support. This work was supported by the National Institutes of Health (R01 GM128439‐01 A1, E.E.C) and the University of Minnesota, Department of Chemistry. N.W.B. was supported by the NIH Institutional Research and Academic Career Development Award K12 GM119955. J.D.S. was supported by the National Institutes of Health's National Center for Advancing Translational Sciences, grants TL1R002493 and UL1TR002494. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health's National Center for Advancing Translational Sciences. Graphical figures were created with BioRender. S. pneumoniae

Funding Information:
The authors thank the M. Winkler Lab at Indiana University for providing S. pneumoniae strains, especially T. Tsui for extensive discussions, and the Carlson Lab for helpful discussion and support. This work was supported by the National Institutes of Health (R01 GM128439-01 A1, E.E.C) and the University of Minnesota, Department of Chemistry. N.W.B. was supported by the NIH Institutional Research and Academic Career Development Award K12 GM119955. J.D.S. was supported by the National Institutes of Health's National Center for Advancing Translational Sciences, grants TL1R002493 and UL1TR002494. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health's National Center for Advancing Translational Sciences. Graphical figures were created with BioRender.

Publisher Copyright:
© 2020 Wiley-VCH GmbH

Keywords

  • activity-based protein profiling
  • bioorthogonal probes
  • click chemistry
  • lactones
  • penicillin-binding proteins

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