No abstract

The grafting-to of a peptide to a side chain functional polymer was investigated using "click" chemistry. Two click reactions were compared: the copper-free strain-promoted azide-alkyne 1,3-cycloaddition (SPAAC) and the traditional copper-catalyzed azide-alkyne 1,3-cycloaddition (CuAAC). For the resulting comb-shaped products, it was found that the steric bulk of the conjugation moiety used in SPAAC limits the degree of grafting for these highly dense systems, whereas CuAAC gives (near) quantitative functionalization.

The cationic amphiphilic antimicrobial peptide gramicidin S (GS) is an effective antibiotic. Its applicability is however restricted to topical infections due to its hemolytic activity. In this study, the process of GS induced hemolysis was investigated in detail for the first time. The morphological changes of red blood cells (RBCs) inflicted by GS were visualized and explained in terms of a physical model. The observed fast rupture events were further investigated with giant unilamellar vesicles (GUVs) as model systems for RBCs. Measurements of membrane fluctuations in GUVs revealed that the membrane surface tension was increased after incubation with GS. These findings are in agreement with the hypothesis that amphiphilic peptides induce membrane rupture by an increase in membrane tension.

The influence of replacing the d-phenylalanine residue with substituted and unsubstituted azoles on the structure and biological activity of the antibiotic gramicidin S was investigated against a representative panel of Gram-positive and Gram-negative bacteria strains. Substituted triazole derivatives, obtained using a convergent synthetic strategy, are as active as gramicidin S, provided that any substituent on the triazole moiety is not too large. The unsubstituted triazole derivative was biologically less active than the parent natural product, gramicidin S. In general for the triazole series, the hemolytic activity could be correlated with the antibacterial activity, that is, the higher the antibacterial activity, the higher the toxicity towards blood cells. Interestingly, its imidazole counterpart showed high antibacterial activity, combined with significantly diminished hemolytic activity.

Synthesis and characterization of peptides 3-17.General methods: (S)-Fmoc-β 3 -homoleucine-OH and (R)-Fmoc-2-aminodecanoic acid were obtained from Senn Chemicals AG. Solvents and chemicals were used as received from their supplier. Solvents were stored over 4 Å molecular sieves. Solvents for column chromatography and extractions were of technical grade and distilled prior to use. THF was distilled over LiAlH 4 , CH 2 Cl 2 over CaH 2 and MeOH over NaBH 4 prior to use. All reactions were performed at room temperature under inert atmosphere. Flash chromatography was performed on Screening Devices silica gel 60 (0.04-0.063 mm). TLC analysis was conducted on DC-alufolien (Merck, Kieselgel 60, F254) with detection by UV-absorption at 254 nm or by spraying with a solution of ninhydrin (3 g/L) in EtOH/AcOH (20:1 v/v), or KMnO 4 solution (20 g in 1% aq K 2 CO 3 ), followed by charring at ~ 150 °C. 1 H and 13 C NMR spectra were recorded with a Bruker DMX-600 spectrometer (600/150 MHz) or with a Bruker AV-400liq spectrometer (400/100 MHz). Chemical shifts (δ) are given in ppm relative to TMS (0.00 ppm) or CD 3 OH (3.31 ppm) as internal standard. CD spectra were recorded at 298 K on a Jasco J-715 spectropolarimeter using 0.2 cm path length quartz cells. The CD spectra are an average of four scans, collected at 0.1 nm intervals between 190 and 250 nm with scanning speed 50 nm/min. The peptides were prepared at concentrations 0.1 mM in MeOH. Ellipticity is reported as mean residue ellipticity [θ], with approximate errors of ±10% at 220 nm. High resolution mass spectra were recorded by direct injection (2 μL of a 2 μM solution in water/acetonitrile; 50:50 v/v and 0.1% formic acid) on a mass spectrometer (Thermo Finnigan LTQ Orbitrap) equipped with an electro spray ion source in positive mode (source voltage 3.5 kV, sheath gas flow 10, capillary temperature 250˚C) with resolution R = 60000 at m/z 400 (mass range m/z = 150-2000) and dioctylphthalate (m/z = 391.28428) as a "lock mass". The high resolution mass spectrometer was calibrated prior to measurements with a calibration mixture (Thermo Finnigan). LC/MS analyses were performed on a LCQ Adventage Max (Thermo Finnigan) equipped with a Gemini C18 column (Phenomenex). The applied buffers were A: H 2 O, B: MeCN and C: 1.0% aq. TFA. HPLC purifications were performed with a Gilson GX-281 automated HPLC system, equipped with a preparative Gemini C18 column (150 × 21.20 mm, 5μ). The applied buffers were: A: 0.2% aq. TFA, B: MeCN. Loading of the HMPB-MBHA-resin:The HMPB-MBHA-resin (theoretical loading = 1.2 mmol/g, 2 mmol, 1.67 g) was suspended in 1,2-dichloroethane (10 mL) and concentrated thrice. Then a solution of the first amino acid (5 eq, 10 mmol), DIC (5 eq, 10 mmol, 1.54 mL) and DMAP (0.01 eq, 20 μmol, 3 mg) in dry DCM/DMF (50 mL, 10:1 v/v) was added. The mixture was shaken for 3 hours and then drained, washed subsequently with DCM, NMP, DCM and Et 2 O. The resin was dried before determination of the loading. The loading procedure was repeated when the loading of the resin wa...