Oxacillin PBP engagement and labeling limits in E. coli DC2
Cell-based study · Live Escherichia coli DC2, an antibiotic-hypersusceptible laboratory mutant; whole-cell competitive fluorescent penicillin labeling. [1]
One coherent whole-cell concentration-response study. Oxacillin preferentially inhibited subsequent Bocillin-FL labeling of PBP3/PBP4. Increased labeling of other targets complicated fitting and limits interpretation of the reported selectivity.
| Endpoint | Reported result | Reference |
|---|---|---|
| Inhibition of subsequent Bocillin-FL labeling of PBP3 and PBP4 | IC50 (estimated) PBP3: 0.09 μg/ml; PBP4: 0.1 μg/ml | [1] |
| Residual Bocillin-FL labeling after 1 μg/ml oxacillin, relative to no-antibiotic control | Relative % Boc-FL labeling PBP3: 36%; PBP4: 34%; averages of two independent experiments | [1] |
| PBP5/6 labeling response at higher oxacillin concentrations | Qualitative fluorescence increase Increased PBP5/6 band intensity; no exact amplitude extracted from graph | [1] |
Literature-Reported Treatment Conditions
- Treatment concentration
- 0.0001 to 1,000 μg/ml oxacillin titration; 1 μg/ml value tabulated separately in Table 1
- Exposure duration
- 30 min oxacillin exposure at room temperature
- Method
- Whole-cell pretreatment, followed by washing and Bocillin-FL labeling; subsequent SDS-PAGE fluorescence analysis relative to no-antibiotic control.
- Vehicle
- Oxacillin stock prepared in purified water; assay exposure in PBS (pH 7.4).
- Reported limits
- Salt/hydration state, product catalog number, lot and lot-specific stereochemical characterization are not reported in the acquired material. SI-only controls were not retrieved; their results are not adopted.
Methods and source locations
Compound reported in the paper: Oxacillin; Compound label in the paper: Oxacillin (Sigma-Aldrich; paper does not specify salt or hydrate)
Oxacillin whole-cell PBP competition results in E. coli DC2 under reported conditions; no salt conversion, universal affinity or clinical efficacy inference.
Tables 1–2 (oxacillin row and footnotes); Figure 2A–C and caption; Materials and Methods: Growth media and antibiotics; β-Lactam titration and detection of PBPs; Results: oxacillin-dependent PBP5/6 labeling
Inhibition of subsequent Bocillin-FL labeling of PBP3 and PBP4: Whole-cell competition followed by SDS-PAGE fluorescence quantification. Four-parameter logistic fitting; Table 2 specifies tenfold concentration steps and estimate values. — Table 2, oxacillin row; footnotes a and c; β-Lactam titration and detection of PBPs
Residual Bocillin-FL labeling after 1 μg/ml oxacillin, relative to no-antibiotic control: Integrated fluorescence band density normalized to untreated samples. Table 1 provides means without a ± term. — Table 1, oxacillin row, column headings and footnotes a–b
PBP5/6 labeling response at higher oxacillin concentrations: Figure 2A gel and 2B quantification; caption identifies standard deviations from two independent experiments. Figure 2C includes untreated and oxacillin-exposed morphology controls. — Figure 2A–C and caption; Results subsection on increased PBP5/6 gel band signal; Table 2 footnote b