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.

Design of multi-functional linear polymers that capture and neutralize a toxic peptide: a comparison with cross-linked nanoparticles.

In this paper, a library of multi-functional linear poly-N-isopropylacrylamide (pNIPAm) polymers having a range of molecular weights and functional groups were synthesized and their interaction with the hemolytic peptide, melittin, was examined. The linear pNIPAm (LPs) containing both tert-butyl group and carboxylic acids bound with the peptide by a combination of hydrophobic and electrostatic interactions and neutralized its toxicity. The melittin binding capacity and affinity of each LP was quantified and further compared with cross-linked multi-functional nanogel particles (NPs) having same combination of functional groups. The binding capacity of the LPs (weight of captured melittin/weight of LP) was independent of their molecular weight and was three times higher than that of previously reported NPs. The binding constant depended on the molecular weight of the LPs, showing the highest value of 1.1 × 108 (M-1) for a ∼1000 mer linear polymer with 40% tert-butyl group and 20% carboxylic acid. Comparison of the interactions of the LPs and NPs suggested the importance of the flexibility of the polymer chain in order to achieve high binding capacity and affinity.

19F-NMR, 1H-NMR, and fluorescence studies of interaction between 5-fluorouracil and polyglycerol dendrimers.

Polyglycerol dendrimers (PGDs), which exhibit a well-defined structure consisting of only glycerol units, were examined as a host molecule of 5-fluorouracil (5-Fu) used as a model anticancer drug. (19)F- and (1)H-NMR titrations and fluorescence measurements were performed to estimate the molecular interaction between PGDs and 5-Fu in a buffer. Results of the NMR titrations revealed that PGD of generation 3 (PGD-G3) encapsulated 5-Fu in the buffer, whereas PGD-G2 and G1 partially incorporated 5-Fu molecule into the space. Fluorescent spectra of 5-Fu in the presence of PGD-G3 indicated that the diketo (lactam) form of 5-Fu changed to the enol-keto (lactim) form of 5-Fu, suggesting attraction of the imine proton of 5-Fu by ether oxygen of PGD-G3. Therefore, the encapsulation state of 5-Fu in PGDs at molecular level was modulated by the well-defined branched structure depending on the generation of PGDs.

Generation-dependent host-guest interactions: solution states of polyglycerol dendrimers of generations†3 and 4 modulate the localization of a guest molecule.

Host-guest interactions between polyglycerol dendrimers of generations 3 and 4 (PGD-G3 and G4) and 4-amino-3-hydroxynapthalene-2-sulphonic acid (AHSA) were investigated by fluorescence spectroscopy, isothermal titration calorimetry (ITC), and dynamic light scattering (DLS). PGD-G3 molecules were found to form an associated state with an average diameter of 82.7 nm in aqueous solution, in which PGD-G3 provided a much more polar microenvironment than glycerol. PGD-G3 and AHSA interacted attractively, showing a binding constant of 5.3×10(5)  M(-1) with a 2:1 stoichiometry. On the other hand, AHSA interacted with the periphery of PGD-G4, the majority of which existed as a unimer, forming a less polar microenvironment. The driving force of the interactions for PGD-G3 and -G4 were mainly enthalpically and entropically driven, respectively. The generation-dependent host-guest interactions were described in conjunction with thermodynamic parameters.

Dendritic nanospace constructed by only glycerol units enhanced uptake of a fluorescent molecule in aqueous solution.

A polyglycerol dendrimer (PGD) of generation 2, which consists of only glycerol units, constructed nanospace capable of uptake of a fluorescent molecule with a 1:1 stoichiometry. On the other hand, a PGD of generation 1 trapped the molecule at the outer part.