Biotin-D-lysine (BDL) and synthetic Lipid II analog were prepared as previously described.29 (link),42 (link) Moenomycin A was isolated from Flavomycin stock. Vancomycin hydrochloride was purchased from Sigma-Aldrich. 2-sulfonatoethyl methanethiosulfonate (MTSES) was purchased from Toronto Research Chemicals. Beta-lactam drugs were purchased from the indicated vendors: piperacillin sodium salt (VWR), imipenem monohydrate (Toronto Research Chemicals), methicillin sodium salt, mecillinam vetranal, cefaclor, oxacillin sodium salt and cephradine (Sigma-Aldrich). S. aureus lipids, 16:0 phosphatidylglycerol (abbreviated as PG), 14:0 cardiolipin (abbreviated as CL), and 16:0 lysyl-phosphatidylglycerol (abbreviated as LPG), were purchased from Avanti Polar Lipids. Nalgene Oak Ridge High-Speed Centrifuge Tubes used for lipid extractions were purchased from Thermo Scientific. Streptavidin-HRP antibody was purchased from Pierce (Catalog #21130). Amersham ECL Prime Western Blotting Detection Reagent was purchased from GE Healthcare. Primers were purchased from Integrated DNA Technologies. Restriction endonucleases were purchased from New England Biolabs. Vectors and expression hosts were obtained from Novagen. Non-stick conical vials and pipette tips used for enzymatic reactions were from VWR. Tryptic Soy Broth and Luria Broth were purchased from Becton Dickinson.
S. aureus strain was grown at 37 °C in Tryptic Soy Broth (TSB) media under aeration with shaking. B. subtilis and E. coli MurJA29C strain were grown at 37 °C in Luria Broth (LB) media under aeration with shaking. LC/MS chromatograms were obtained on an Agilent Technologies 1100 series LC-MSD instrument using electrospray ionization (ESI). HRMS data was obtained on a Bruker Maxis Impact LC-q-TOF Mass Spectrometer using ESI. Western blots were developed using Biomax Light Film (Kodak) or imaged using an Azure C400 imaging system. ImageJ was used for densitometric analysis of western blots.
S. aureus strain was grown at 37 °C in Tryptic Soy Broth (TSB) media under aeration with shaking. B. subtilis and E. coli MurJA29C strain were grown at 37 °C in Luria Broth (LB) media under aeration with shaking. LC/MS chromatograms were obtained on an Agilent Technologies 1100 series LC-MSD instrument using electrospray ionization (ESI). HRMS data was obtained on a Bruker Maxis Impact LC-q-TOF Mass Spectrometer using ESI. Western blots were developed using Biomax Light Film (Kodak) or imaged using an Azure C400 imaging system. ImageJ was used for densitometric analysis of western blots.
