Ethyl hydroxyimino cyanoacetate

A CEM microwave synthesizer (Liberty Blue) was used for solid phase peptide synthesis of PTs, as previously described [14 (link), 21 (link), 22 (link), 39 (link)]. Fmoc-4-azido-Proline used in the synthesis was prepared as previously reported [21 (link)]. All other Fmoc-, Boc- protected amino acids, N,Nʹ-diisopropylcarbodiimide, ethyl (hydroxyimino) cyanoacetate (OxymaPure) and rink amide resin (100–200 mesh size, 0.53 meq/g substitution) were purchased from Chem-Impex International, INC. Ethynylferrocene and CuI catalyst for the click reaction, and hydrazine, were purchased from Sigma Aldrich. HPLC purifications were performed using a Waters® HPLC system with reverse phase (RP) C18 prep columns. Purity verification of cPTs was carried out by analytical C18 RP-HPLC column, using a BeckmanCoulter® HPLC system. HPLC grade acetonitrile (ACN), Millipore-MilliQ water and 0.1% TFA were used as solvents for the HPLC purification. Mass was confirmed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS).

The bacterial strains (methicillin-sensitive S. aureus (MSSA) ATCC 25923, methicillin-resistant Staphylococcus aureus (MRSA) ATCC43300, E. coli ATCC 25922, and P. aeruginosa ATCC 27853) were obtained from the American Type Culture Collection (Manassas, VA, USA). All amino acids and Rink amide-4-methylbenzhydrylamine (MBHA) resin (0.54 mmol/g) were obtained from Mimotopes (Melbourne, VIC, Australia). Ethyl (hydroxyimino)cyanoacetate (Oxyma Pure) and N,N′-diisopropylcarbodiimide (DIC) were obtained from Chem-Impex International (Wood Dale, IL, USA). Triisopropylsilane (TIPS) was obtained from Alfa Aesar (Massachusetts, VA, USA). Dulbecco’s Modified Eagle Medium (DMEM), fetal bovine serum (FBS), and penicillin/streptomycin were obtained from Thermo Fisher Scientific (Scoresby, VIC, Australia). Acetonitrile (MeCN), cetyltrimethylammonium bromide (CTAB, 99%), N,N-dimethylformamide (DMF; peptide synthesis grade), dimethyl sulfoxide (DMSO), ethanol (EtOH), methanol (MeOH), Mueller–Hinton broth (MHB), tetraethyl orthosilicate (TEOS, reagent grade 98%), trifluoroacetic acid (TFA), 3-(trihydroxysilyl)propyl methylphosphonate (THMP, 90%), and all other reagents were purchased from Merck (Castle Hill, NSW, Australia).