Selective penicillin-binding protein imaging probes reveal substructure in bacterial cell division

Ozden Kocaoglu, Rebecca A. Calvo, Lok To Sham, Loralyn M. Cozy, Bryan R. Lanning, Samson Francis, Malcolm E. Winkler, Daniel B. Kearns, Erin E. Carlson

Research output: Contribution to journalArticlepeer-review

56 Scopus citations

Abstract

The peptidoglycan cell wall is a common target for antibiotic therapy, but its structure and assembly are only partially understood. Peptidoglycan synthesis requires a suite of penicillin-binding proteins (PBPs), the individual roles of which are difficult to determine because each enzyme is often dispensable for growth perhaps due to functional redundancy. To address this challenge, we sought to generate tools that would enable selective examination of a subset of PBPs. We designed and synthesized fluorescent and biotin derivatives of the β-lactam-containing antibiotic cephalosporin C. These probes facilitated specific in vivo labeling of active PBPs in both Bacillus subtilis PY79 and an unencapsulated derivative of D39 Streptococcus pneumoniae. Microscopy and gel-based analysis indicated that the cephalosporin C-based probes are more selective than BOCILLIN-FL, a commercially available penicillin V analogue, which labels all PBPs. Dual labeling of live cells performed by saturation of cephalosporin C-susceptible PBPs followed by tagging of the remaining PBP population with BOCILLIN-FL demonstrated that the two sets of PBPs are not co-localized. This suggests that even PBPs that are located at a particular site (e.g., septum) are not all intermixed, but rather that PBP subpopulations are discretely localized. Accordingly, the Ceph C probes represent new tools to explore a subset of PBPs and have the potential to facilitate a deeper understand of the roles of this critical class of proteins.

Original languageEnglish (US)
Pages (from-to)1746-1753
Number of pages8
JournalACS Chemical Biology
Volume7
Issue number10
DOIs
StatePublished - Oct 19 2012

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