Stimuli-Responsive Delivery of Antimicrobial Peptides Using Polyelectrolyte Complexes.

Antimicrobial peptides (AMPs) are antibiotics with the potential to address antimicrobial resistance. However, their translation to the clinic is hampered by issues such as off-target toxicity and low stability in biological media. Stimuli-responsive delivery from polyelectrolyte complexes offers a simple avenue to address these limitations, wherein delivery is triggered by changes occurring during microbial infection. The review first provides an overview of pH-responsive delivery, which exploits the intrinsic pH-responsive nature of polyelectrolytes as a mechanism to deliver these antimicrobials. The examples included illustrate the challenges faced when developing these systems, in particular balancing antimicrobial efficacy and stability, and the potential of this approach to prepare switchable surfaces or nanoparticles for intracellular delivery. The review subsequently highlights the use of other stimuli associated with microbial infection, such as the expression of degrading enzymes or changes in temperature. Polyelectrolyte complexes with dual stimuli-response based on pH and temperature are also discussed. Finally, the review presents a summary and an outlook of the challenges and opportunities faced by this field. This review is expected to encourage researchers to develop stimuli-responsive polyelectrolyte complexes that increase the stability of AMPs while providing targeted delivery, and thereby facilitate the translation of these antimicrobials.

Nisin:Carboxymethylcellulose polyion complex (PIC) nanoparticles. Preparation and antimicrobial activity.

Nisin is a cationic antimicrobial peptide used as a natural food preservative against gram-positive bacteria. However, nisin is degraded following interaction with food components. Here, we report the first use of Carboxymethylcellulose (CMC), a versatile and affordable food additive, to protect nisin and extend its antimicrobial activity. First, we optimized the methodology by considering the effect of nisin:CMC ratio, pH, and, especially, the degree of substitution of CMC. In particular, we show here how these parameters affected the size, charge, and, notably, the encapsulation efficiency of these nanomaterials. This way, optimized formulations contained over 60 % w/w in nisin while encapsulating ∼90 % of the nisin used. We then show that these new nanomaterials inhibited the growth of Staphylococcus aureus, a major foodborne pathogen, using milk as a representative food matrix. Remarkably, this inhibitory effect was observed with one-tenth of the concentration of nisin currently used in dairy products. We believe that the combination of the affordability of CMC, flexibility and simplicity of preparation, and the ability to inhibit the growth of food pathogens, makes these nisin:CMC PIC nanoparticles an ideal platform to underpin new nisin formulations.

Polymer-induced biofilms for enhanced biocatalysis.

The intrinsic resilience of biofilms to environmental conditions makes them an attractive platform for biocatalysis, bioremediation, agriculture or consumer health. However, one of the main challenges in these areas is that beneficial bacteria are not necessarily good at biofilm formation. Currently, this problem is solved by genetic engineering or experimental evolution, techniques that can be costly and time consuming, require expertise in molecular biology and/or microbiology and, more importantly, are not suitable for all types of microorganisms or applications. Here we show that synthetic polymers can be used as an alternative, working as simple additives to nucleate the formation of biofilms. Using a combination of controlled radical polymerization and dynamic covalent chemistry, we prepare a set of synthetic polymers carrying mildly cationic, aromatic, heteroaromatic or aliphatic moieties. We then demonstrate that hydrophobic polymers induce clustering and promote biofilm formation in MC4100, a strain of Escherichia coli that forms biofilms poorly, with aromatic and heteroaromatic moieties leading to the best performing polymers. Moreover, we compare the effect of the polymers on MC4100 against PHL644, an E. coli strain that forms biofilms well due to a single point mutation which increases expression of the adhesin curli. In the presence of selected polymers, MC4100 can reach levels of biomass production and curli expression similar or higher than PHL644, demonstrating that synthetic polymers promote similar changes in microbial physiology than those introduced following genetic modification. Finally, we demonstrate that these polymers can be used to improve the performance of MC4100 biofilms in the biocatalytic transformation of 5-fluoroindole into 5-fluorotryptophan. Our results show that incubation with these synthetic polymers helps MC4100 match and even outperform PHL644 in this biotransformation, demonstrating that synthetic polymers can underpin the development of beneficial applications of biofilms.

Morphological transition and transformation of 2D nanosheets by controlling the balance of π-π stacking interaction and crystalline driving forces.

Nanoscale organic two-dimensional (2D) materials of block polymers (BCPs) have attracted interest on account of their wide potential applications in a range of fields. Herein, we design a new poly(p-phenylenevinylene) (PPV) based BCP that contains a triisopropylsilyl side chain and poly (2-vinyl pyridine) (P2VP) corona, which could assemble into a series of 2D square and rectangular micelles in isopropanol. The aspect ratios and the scales of the 2D micelles can be tuned in two ways, including altering the ratios of the P2VP and PPV-TIPS blocks and their concentrations. By precisely controlling the aspect ratios, micro-scale rod-like micelles are also obtained. From in depth studies of the morphology transition from rectangular micelles to rod-like or square micelles, it is found that the BCPs initially organize into fibers and then assemble into final micelles by the combined forces of π-π interactions and the crystalline force from TIPS side chains. Based on the balance of the two interactions, 2D circle-like micelles are also achieved by heterogenous co-assembly of two kinds of polymers with different cores.

Morphologically Tunable Supramolecular Rectangular Microsheet and Microsaw Constructed by Hierarchical Self-Assembly Based on Hydrogen Bonds.

An amino acid derivative, thiophene (TDAV), as new building blocks for 2D supramolecular assembly is designed. Various square and rectangular microsheets are achieved and the aspect ratios are precisely regulated by controlling the polarity of cosolvent or water content. By the introduction of chirality, the novel microsaw is also achieved. It provides a new approach to prepare various kinds of unique supramolecular 2D materials with controllable shapes and sizes for future biological applications.

Hierarchical Chiral Supramolecular Nanoarchitectonics with Molecular Detection: Helical Structure Controls upon Self-Assembly and Coassembly.

The morphological transformation from microspheres to helical supramolecular nanofibers with controllable handedness is achieved by the introduction of molecular chirality based on amino acid derivatives (TDAP), and the chirality of the supramolecular architectures that are achieved is nullified through the coassembly of the equivalent TDAP enantiomers. The molecular detection of achiral melamine based on the R-TDAP-COOH supramolecular system is achieved by the appearance of helicity and inversion.

Site-Selective and Rewritable Labeling of DNA through Enzymatic, Reversible, and Click Chemistries.

Current methods for bioconjugation rely on the introduction of stable linkers that lack the required versatility to perform sequential functionalizations. However, sequential manipulations are an increasing requirement in chemical biology because they can underpin multiple analyses of the same sample to provide a wider understanding of cell behavior. Here, we present a new method to site-selectively write, remove, and rewrite chemical functionality to a biomolecule, DNA in this case. Our method combines the precision and robustness of methyltransferase-directed labeling with the reversibility of acyl hydrazones and the efficiency of click chemistry. Underpinning the method is a new S-adenosyl-l-methionine derivative to site-selectively label DNA with a bifunctional chemical handle containing an acyl hydrazone-linker and a terminal azide. Functional tags are conjugated via the azide and can be removed (i.e., untagged) when needed at the acyl hydrazone via exchange with hydroxyl amine. The formed hydrazide-labeled DNA is a versatile intermediate that can be either rewritten to reset the original chemical handle or covalently reacted with a permanent tag. This ability to write, tag, untag, and permanently tag DNA is exploited to sequentially introduce two fluorescent dyes on DNA. Finally, we demonstrate the potential of the method by developing a protocol to sort labeled DNA using magnetic beads, with subsequent amplification of the sorted DNA sample for further analysis. The presented method opens new avenues for site-selective bioconjugation and should underpin integrative approaches in chemical biology where sequential functionalizations of the same sample are required.

Helical polymers for biological and medical applications.

Helices are the most prevalent secondary structure in biomolecules and play vital roles in their activity. Chemists have been fascinated with mimicking this molecular conformation with synthetic materials. Research has now been devoted to the synthesis and characterization of helical materials, and to understand the design principles behind this molecular architecture. In parallel, work has been done to develop synthetic polymers for biological and medical applications. We now have access to materials with controlled size, molecular conformation, multivalency or functionality. As a result, synthetic polymers are being investigated in areas such as drug and gene delivery, tissue engineering, imaging and sensing, or as polymer therapeutics. Here, we provide a critical view of where these two fields, helical polymers and polymers for biological and medical applications, overlap. We have selected relevant polymer families and examples to illustrate the range of applications that can be targeted and the impact of the helical conformation on the performance. For each family of polymers, we briefly describe how they can be prepared, what helical conformations are observed and what parameters control helicity. We close this Review with an outlook of the challenges ahead, including the characterization of helicity through the process and the identification of biocompatibility.

Messenger RNA delivery by hydrazone-activated polymers.

The intracellular delivery of DNA and RNA therapeutics requires the assistance of vectors and/or nucleotide modifications to protect the nucleic acids against host nucleases and promote cellular internalization and release. Recently, messenger RNA (mRNA) has attracted much attention due to its transient activity and lack of genome permanent recombination and persistent expression. Therefore, there is a strong interest in the development of conceptually new non-viral vectors with low toxicity that could improve mRNA transfection efficiency. We have recently introduced the potential of polyhydrazones and the importance of the degree of polymerization for the delivery of siRNA and plasmid DNA. Here, we demonstrate that this technology can be easily adapted to the more interesting complexation and delivery inside living cells of mRNA. The polyplexes resulting from the combination of the amphiphilic polyhydrazone were characterized and the transfection efficiency and cell viability were studied for a discrete collection of functionalized polyhydrazones. The results obtained demonstrated the versatility of these polymeric vectors as excellent candidates for the delivery of mRNA and validate the easy adaptability of the technology to more sensitive and therapeutically relevant nucleic acids.