A Thermoresponsive Cationic Comb-Type Copolymer Enhances Membrane Disruption Activity of an Amphiphilic Peptide

Membrane active peptides (MAPs) have potential applications in drug delivery systems and as antimicrobials. We previously showed that a cationic comb-type copolymer, poly­(allylamine)-<i>graft</i>-dextran (PAA-<i>g</i>-Dex), forms a soluble inter-polyelectrolyte complex with an anionic MAP, the E5 peptide, resulting in significant enhancement of the membrane disruption activity of E5. In this study, we designed a novel comb-type cationic copolymer composed of a PAA main chain and thermoresponsive poly­(<i>N</i>-isopropyl­acrylamide) graft chains (PAA-<i>g</i>-PNIPAAm). We hypothesized that the thermoresponsive hydrophilic/hydrophobic transition of the grafted polymer would regulate the membrane disruption activity of E5 peptide. Both the binding affinity of the complex and the membrane disruption activity of E5/PAA-<i>g</i>-PNIPAAm were found to be enhanced above the phase transition temperature of the grafted chain. Our analysis suggests that the hydrophilic/hydrophobic environment around the cationic polymer chain plays important roles in the enhancement of the activity of the anionic peptide

Design of UCST Polymers for Chilling Capture of Proteins

Ureido-derivatized polymers, such as poly­(allylurea) (PU) and poly­(_L-citrulline) derivatives, exhibited upper critical solution temperature (UCST) behavior under physiological buffer conditions as we previously reported. The PU derivatives having amino groups (PU-Am) also showed UCST behavior. In this study, we modified the amino groups of the polymer with succinyl anhydride (PU-Su) or acetyl anhydride (PU-Ac) to determine the effects of these ionic groups on the UCST behavior and to control interactions between the PU derivatives and biocomponents such as proteins and cells. Succinylation of PU-Am resulted in a significant decrease in phase separation temperature (<i>T</i>_p), whereas acetylation of PU-Am resulted in an increase in <i>T</i>_p. As expected, the <i>T</i>_p of PU-Am and PU-Su changed when the pH of the solution was changed. The <i>T</i>_p of PU-Am increased at higher pH, whereas that of PU-Su increased at lower pH, indicating that ionic charge decreases <i>T</i>_p of PU derivatives by increasing osmotic pressure and by increasing hydrophilicity of the polymer chains. Interestingly, these groups did not significantly change UCST when these groups were nonionic. We then examined capture and separation of particular proteins from a protein mixture by cooling-induced phase separation. Selective and rapid capture of particular proteins from protein mixture by PU derivatives was shown, indicating that the ureido-derivatized polymers are potential media for bioseparation under biofriendly conditions

All data underlying the findings

Data for 3 figures (sheets 1-3) and qPCR results for 6 plates (sheets 7-12

Time-dependent changes in the eDNA concentration after fish removal.

<p>Regression curves of the non-linear models are shown. Each color indicates one of five individuals (black and aqua: adult, others: juvenile).</p

Initial eDNA concentration and degradation constant (<i>N</i>₀ and <i>ß</i> respectively; ±SE) estimated by non-linear models fitted to the change in the eDNA concentration after fish removal and fish body wet weight.

<p>Significance levels (<i>t</i>-test) are indicated by *** (<i>p</i><0.001), ** (<i>p</i><0.01), and * (<i>p</i><0.05).</p><p>Initial eDNA concentration and degradation constant (<i>N</i>₀ and <i>ß</i> respectively; ±SE) estimated by non-linear models fitted to the change in the eDNA concentration after fish removal and fish body wet weight.</p

Time-dependent changes in the eDNA concentration after fish introduction.

<p>Plots and error bars indicate means and standard deviations of five individuals, respectively.</p

Box plots of the eDNA release compared between juvenile and adult groups.

<p>a) Stabilized concentration, b) release rate per individual fish, and c) per fish body weight. Body wet weight was 0.5–2.0 g (<i>n</i> = 10) and 30–75 g (<i>n</i> = 9), respectively.</p

Minimization of Synthetic Polymer Ligands for Specific Recognition and Neutralization of a Toxic Peptide

Synthetic polymer ligands (PLs) that recognize and neutralize specific biomacromolecules have attracted attention as stable substitutes for ligands such as antibodies and aptamers. PLs have been reported to strongly interact with target proteins and can be prepared by optimizing the combination and relative proportion of functional groups, by molecular imprinting polymerization, and/or by affinity purification. However, little has been reported about a strategy to prepare PLs capable of specifically recognizing a peptide from a group of targets with similar molecular weight and amino acid composition. In this study, we show that such PLs can be prepared by minimization of molecular weight and density of functional units. The resulting PLs recognize the target toxin exclusively and with 100-fold stronger affinity from a mixture of similar toxins. The target toxin is neutralized as a result. We believe that the minimization approach will become a valuable tool to prepare “plastic aptamers” with strong affinity for specific target peptides

Media 3: Wide field intravital imaging by two-photon-excitation digital-scanned light-sheet microscopy (2p-DSLM) with a high-pulse energy laser

Originally published in Biomedical Optics Express on 01 October 2014 (boe-5-10-3311

Media 1: Wide field intravital imaging by two-photon-excitation digital-scanned light-sheet microscopy (2p-DSLM) with a high-pulse energy laser

Originally published in Biomedical Optics Express on 01 October 2014 (boe-5-10-3311