Article Figures & Data
Figures
- Fig. 1.
CD spectra of cationic peptides in the presence of 90-nm liposomes (POPC-POPG, 7:3) and CP26 in phosphate buffer (random coil). The peptides are represented as follows: CP201, round dots; CP208, solid lines; CEMA, dash-dots; CEME, long dashes; CP26, dashes; and CP29, square dots.
- Fig. 2.
Survival of logarithmic-phase (A) and stationary-phase (B) E. coli UB1005 in MH broth after addition of fourfold the MIC of peptide, gentamicin, or ceftazidime. This corresponds to 2 μg of CP26, CP29, and ceftazadime per ml, 4 μg of CEME and CEMA per ml, and 0.5 μg of gentamicin per ml. Actual initial concentrations of bacteria ranged from 0.5 × 108 to 2.5 × 108/ml but were corrected to an initial concentration of 1 × 108/ml for clarity. A typical experiment out of three trials is shown. Symbols: ▵, no peptide; ▴, CP26; ▾, CP29; ●, CEME; ○, CEMA; ■, ceftazidime; □, gentamicin. Results for CP26 in panel A were almost superimposable with the ceftazidime results and were thus omitted for clarity. No evidence of bacterial aggregation was observed when viewed under a light microscope or in light-scattering experiments.
- Fig. 3.
Permeabilization of cytoplasmic membrane of E. coli ML35 by cationic peptides as determined by their ability to unmask cytoplasmic β-galactosidase. The hydrolysis of ONPG was measured spectrophotometrically at 420 nm. Symbols: ▴, CP26 at 12.8 μg/ml; ●, CEME at 6.4 μg/ml; ○, CEMA at 6.4 μg/ml; ▾, CP29 at 6.4 μg/ml; ■, polymyxin B at 12.8 μg/ml; □, CP201 at 12.8 μg/ml; ▵, CP208 at 12.8 μg/ml.
- Fig. 4.
Effects of NaCl, MgCl2, and CCCP on the cytoplasmic membrane permeabilization activity of cationic peptides CP29 (solid), CEME (stippled), and CP26 (striped).
Tables
- Table 1.
Amino acid sequences of antimicrobial cationic peptides used in this study
Peptide Amino acid sequence Length (no. of amino acids) Charge % Hydrophobic amino acids CP26 KWKSFIKKLTSAAKKVVTTAKPLISS 26 +7 46 CP29 KWKSFIKKLTTAVKKVLTTGLPALIS 26 +6 50 CP201 KWKSFIKNLTKGGSKILTTGLPALIS 26 +5 42 CP208 KKKSFIKLLTSAKVSVLTTAKPLISS 26 +6 42 CEME KWKLFKKIGIGAVLKVLTTGLPALIS 26 +5 69 CEMA KWKLFKKIGIGAVLKVLTTGLPALKLTK 28 +6 64 - Table 2.
α-Helicity in various environments as assessed by CD spectroscopy interpreted according to the K2D algorithm (2)
Condition % α-Helix CP26 CP29 CP201 CP208 CEMA CEME Phosphate buffer 6 8 8 7 4 7 Liposomesa 26, 35 50, 57 20 8 17, 33 29, 30 SDS (40:1) 42 23 13 26 50 35 50% TFE 30 20 ND ND 32 25 ↵a The first number is for 90-nm liposomes, and the second is for 60-nm liposomes. ND, not determined.
- Table 3.
Broth dilution MICs of cationic peptides against various gram-negative bacteria
Species Strain Relevant phenotype or genotype MIC (μg/ml) CP26 CP29 CP201 CP208 CEME CEMA P. aeruginosa PAO1 Wild type 2 2 8 64 2 2 K385 nfxB 4 4 8 64 2 2 1008OCR01 nalB 2 2 16 128 2 2 PAO963 nalA 2 2 8 128 2 2 PA-83-48 β-Lactamase derepressed 4 2 32 >128 2 4 Z61 Antibiotic sensitive 2 2 32 128 2 4 E. coli UB1005 Wild type 0.5 0.5 4 32 1 1 S. typhimurium 14028s Parent of MS7953s 2 2 32 >64 2 4 MS7953s Defensin sensitive 0.5 0.25 2 8 1 1 - Table 4.
Influence of NaCl, MgCl2, and polyanionic alginate on MICs of cationic antimicrobial peptides forP. aeruginosa PAO1
Additive Concn MIC (μg/ml)a CP26 CP29 CEME CEMA None 2 2 2 2 NaCl 100 mM 4 2 2 2 200 mM 16 2 4 4 300 mM 32 4 4 4 MgCl2 1 mM 8 4 4 4 3 mM 16 16 16 4 5 mM >32 32 32 32 Sodium alginate 0.01% 2 2 4 4 0.05% 8 8 16 32 0.1% 16 32 64 64 ↵a These are the average values of three experiments.
