9 research outputs found
Amphiphilic Cationic β<sup>3R3</sup>-Peptides: Membrane Active Peptidomimetics and Their Potential as Antimicrobial Agents
We introduce a novel
class of membrane active peptidomimetics,
the amphiphilic cationic β<sup>3R3</sup>-peptides, and evaluate
their potential as antimicrobial agents. The design criteria, the
building block and oligomer synthesis as well as a detailed structure–activity
relationship (SAR) study are reported. Specifically, infrared reflection
absorption spectroscopy (IRRAS) was employed to investigate structural
features of amphiphilic cationic β<sup>3R3</sup>-peptide sequences
at the hydrophobic/hydrophilic air/liquid interface. Furthermore,
Langmuir monolayers of anionic and zwitterionic phospholipids have
been used to model the interactions of amphiphilic cationic β<sup>3R3</sup>-peptides with prokaryotic and eukaryotic cellular membranes
in order to predict their membrane selectivity and elucidate their
mechanism of action. Lastly, antimicrobial activity was tested against
Gram-positive M. luteus and S. aureus as well as against Gram-negative E. coli and P. aeruginosa bacteria along with testing hemolytic activity and cytotoxicity.
We found that amphiphilic cationic β<sup>3R3</sup>-peptide sequences
combine high and selective antimicrobial activity with exceptionally
low cytotoxicity in comparison to values reported in the literature.
Overall, this study provides further insights into the SAR of antimicrobial
peptides and peptidomimetics and indicates that amphiphilic cationic
β<sup>3R3</sup>-peptides are strong candidates for further development
as antimicrobial agents with high therapeutic index
Magnetic Porous Sugar-Functionalized PEG Microgels for Efficient Isolation and Removal of Bacteria from Solution
Here,
we present a new microparticle system for the selective detection
and magnetic removal of bacteria from contaminated solutions. The
novelty of this system lies in the combination of a biocompatible
scaffold reducing unspecific interactions with high capacity for bacteria
binding. We apply highly porous poly(ethylene glycol) (PEG) microparticles
and functionalize them, introducing both sugar ligands for specific
bacteria targeting and cationic moieties for electrostatic loading
of superparamagnetic iron oxide nanoparticles. The resulting magnetic,
porous, sugar-functionalized (MaPoS) PEG microgels are able to selectively
bind and discriminate between different strains of bacteria Escherichia coli. Furthermore, they allow for a highly
efficient removal of bacteria from solution as their increased surface
area can bind three times more bacteria than nonporous particles.
All in all, MaPoS particles represent a novel generation of magnetic
beads introducing for the first time a porous, biocompatible and easy
to functionalize scaffold and show great potential for various biotechnological
applications
GPIs of RH and PTG strains induce TNF-α and IL-12p40 secretion by macrophages.
<p>(<b>A</b>) Macrophages were incubated for 24 h with medium alone, or with individual GPIs of the PTG strain extracted from 1×10<sup>8</sup> parasites and assayed for TNF-α cytokine production. (<b>B</b>) Macrophages were incubated for 24 h with medium alone, or with individual GPIs of the RH (left panel) and the PTG strain (right panel) extracted from 2×10<sup>8</sup> parasites, respectively, and assayed for IL-12p40 cytokine production. (<b>C</b>) Macrophages were incubated for 24 h with medium alone, or with GPIa (3 mM), a chemically synthesized structure of RH strain GPI III core glycan and assayed for IL-12p40 cytokine production. <sup>***</sup>P<0.0005 PTG GPI II compared to medium control. ND, not determined.</p
Protein-free Glc-GalNAc-substituted GPIs are clustered on extracellular parasites.
<p>Staining after permeabilization of HFF cells infected (72 h p.i.) with RH (upper panel) and PTG (lower panel) strains was performed using mAb T54 E10, recognizing both the EtN-PO<sub>4</sub> and the Glc-GalNAc side-branch epitopes of protein-free GPIs. Filled arrowheads point to intracellular parasites residing inside parasitophorous vacuoles. Unfilled arrowheads point to extracellular parasites. DIC, differential interference contrast.</p
Comparison of AHM and inositol quantifications of protein-free GPIs and GPI-anchored proteins of RH and PTG strains.
a<p>Copies per cell. In parentheses are percentages of PTG values compared to RH values. Values are means ± SD of three replicates.</p
GPIs of the PTG strain activate TLR4/NF-κB signaling.
<p>(<b>A</b>) CHO cells expressing TLR2 (TLR2<sup>+</sup>), TLR4 (TLR4<sup>+</sup>), or neither (TLR2<sup>−</sup>/TLR4<sup>−</sup>) were either untreated (black line) or exposed to the four different GPIs (II, III, V and VI) extracted from 1×10<sup>9</sup> parasites (gray line). CD25 expression was measured by FACS analysis 18 h after stimulation. The figure is representative of three independent experiments. Percentage = [percentage of CD25 expression (M2) on GPI-stimulated cells] minus [percentage of CD25 expression (M2) on medium stimulated cells]. (<b>B</b>) Macrophages were incubated for 15, 30, or 60 min with GPI VI extracted from 4×10<sup>8</sup> PTG strain parasites. Total nuclear protein content was tested using a NF-κB assay as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0085386#s2" target="_blank">Materials and Methods</a>. The figure is a representative experiment with PTG GPI VI and similar results were obtained with GPIs II, III and V of the PTG strain. <sup>**</sup>P<0.005 compared to medium control.</p
RH and PTG strains have different GPI profiles.
<p>(<b>A</b>) Metabolically labeled ([<sup>3</sup>H]-glucosamine) parasite glycolipids were extracted and separated by TLC as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0085386#s2" target="_blank">Materials and Methods</a>. TLC chromatograms were scanned for radioactivity using a Berthold LB 2842 linear analyzer. Six different peaks representing individual GPIs are detected in the RH strain <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0085386#pone.0085386-Striepen1" target="_blank">[35]</a>, whereas only four major peaks are present in the PTG strain. (<b>B</b>) Structures of <i>T. gondii</i> RH GPIs. GPI VI (Man-Man-[GalNAc]Man-GlcN-PI) is neither substituted with an EtN-PO<sub>4</sub> nor with the Glc linked to the GalNAc residue. GPIs I, II and III contain an EtN-PO<sub>4</sub> residue (dashed frame line). GPIs I, II, IV and V are substituted with a Glc that is linked to the GalNAc side-branch in α1–4 linkage (thickened frame line). GPIs I, II and IV, V, respectively, have identical carbohydrate moieties and differ in their fatty acid composition.</p
GPI core glycans and PIs of RH and PTG strains have similar structures.
<p>(<b>A</b>) HPAEC analysis of the core glycans generated from <i>T. gondii</i> PTG strain protein-free GPIs II, III, V and VI. TLC-purified D-[<sup>3</sup>H]-glucosamine labeled GPIs were dephosphorylated, deaminated and reduced. The resulting neutral glycans were analysed before (untreated) and after Jack bean α-mannosidase or hexosaminidase treatments. The elution positions of the co-injected glucose oligomer standards are indicated at the top of each profile (bold digits) and given as DU. (<b>B</b>) Predicted Man-Man-(Glc-GalNAc)Man-anhydromannitol core glycan structure derived from protein-free GPIs II and V (right panel), and predicted Man-Man-(GalNAc)Man-anhydromannitol core glycan structure derived from protein-free GPIs III and VI (left panel) of the PTG strain. (<b>C</b>) ES-MS spectra of the PI moieties released by deamination from purified protein-free GPIs from RH (upper panel) and PTG (lower panel) strains.</p
Carbohydrate composition analysis of protein-free GPIs and GPI-anchored proteins of RH and PTG strains.
a<p>Glucose is a common contaminant. Values are means of molar ratios ± SD of three replicates.</p><p>Values are means of molar ratios ± SD of three replicates.</p>**<p>P<0.005,</p>*<p>P<0.05 RH protein-free GPI and GPI-anchored protein samples compared to PTG protein-free GPI and GPI-anchored protein samples, respectively.</p