cereus and Gram-negative E. coli were incubated with increasing concentrations of mono-PEG-StAP3 fraction for 6 h at 37 °C. Results obtained here show that mono-PEG-StAP3 was able to kill bacterial cells in a dose-dependent manner ( Fig. 4). The antibacterial activity of mono-PEG-StAP3 was more effective against B. cereus than E. coli. The IC50 values were approximately 13.2 and 96.2 μg/ml mono-PEG-StAP3, respectively ( Table 1). The IC50 values of mono-PEG-StAP3 Afatinib nmr were approximately 4 times

lower on B. cereus, and approximately 1.6 times higher on E. coli compared to the StAP3 native form [30]. The greater susceptibility of mono-PEG-StAP3′s antimicrobial effect on B. cereus compared to E. coli may be accounted for the bacterial cell membrane

composition. Gram-negative bacteria have a cytoplasmatic membrane and an additional outer membrane that surrounds the cell, providing a barrier to mono-PEG-StAP3, whereas Gram-positive bacteria have only cytoplasmatic membrane [65] and [66]. In comparison, PEGylation of antimicrobial peptides Epacadostat tachyplesin I, nisin, α-defensin, and magainin with 5 kDa PEG chains led to a drastic decrease or even a complete loss of their antibacterial activities [64], [67], [68] and [69]. Nevertheless, the extent of the reduction in activity is strongly dependent on the peptide/protein evaluated. It is possible that mono-PEG-StAP3 decreases its ability to efficiently permeate the outer membrane due

to a large steric hindrance of the PEG moiety, similar to that reported for PEGylated tachyplesin I and magainin [64] and [68]. Some antimicrobial peptides such as melittin, gramicidin S, CaLL, and surfactant protein B are also cytotoxic to mammalian cells, e.g. erythrocytes [70], [71], [72] and [73]. Therefore, only antimicrobial peptides/proteins and their derivatives with high antimicrobial activity and low cytotoxicity to the healthy eukaryotic cells are of practical interest. The hemolytic activity of mono-PEG-StAP3 fraction was tested in vitro on hRBC to investigate whether PEGylation affects the selective cytotoxicity of StAP3. As shown in Table 2, mono-PEG-StAP3 did not show significant hemolytic activity at all concentrations assayed. Several reports relate the hemolytic activity of antimicrobial peptides with their capacity to Cediranib (AZD2171) strongly interact with either membranes, containing cholesterol or not [74] and [75]. As for the case of antimicrobial peptides unable to lyse red blood cells [76], the presence of cholesterol into the LUVs membranes strongly diminishes the capacity of StAsp-PSI to produce leakage at all concentration assayed [29]. The presence of cholesterol in the membranes causes a reduction in the density of hydrophilic head groups at the interfacial region of the bilayer and an increase in the packaging of the phospholipid tails in the middle of the bilayer [77].