Modular Ligation of Thioamide Functional Peptides onto Solid Cellulose Substrates

Hetero Diels‐Alder (HDA) cycloaddition – as an effective modular conjugation approach – is employed to graft thioamide endfunctional oligopeptides onto solid cyclopentadienyl (Cp) functional cellulose substrates generating cellulose‐peptide hybrid materials. The highly reactive Cp moieties serve as diene functionality in the consecutive HDA reaction on the biosubstrate surface. Oligopeptides (i.e., the model peptide Gly‐Gly‐Arg‐Phe‐Pro‐Trp‐Trp‐Gly and the antimicrobial peptide tritrpticin) are functionalized at their N‐termini employing strongly electron deficient thiocarbonyl thio compounds resulting in biomacromolecules bearing a thioamide endgroup. The dienophile‐ functional peptides readily undergo HDA reactions at ambient temperature and under mild conditions in solution with synthetic polymers as well as on solid (bio)substrates. An in‐depth investigation is provided of the influence of the temperature, the Lewis acid catalysis and the side group exchange of thioamide functional oligopeptides reacting with Cp terminated poly(methyl methacrylate) (M_n = 2100 g·mol^?1, PDI = 1.1) in homogenous solution as well as Cp functionalized cellulose in a heterogeneous system. To assess the success of the grafting reaction, the soluble samples were subjected to characterization methods such as size exclusion chromatography (SEC) and SEC‐electrospray ionization mass spectrometry (SEC‐ESI‐MS). The heterogeneous “grafting‐to” reactions were monitored using high resolution attenuated total reflection (ATR) Fourier transform infrared microscopy (HR‐FTIRM) imaging, X‐ray photoelectron spectroscopy (XPS) and elemental analysis. Evaluation via elemental analysis leads to quantitative peptide cellulose surface loading capacities.     

Hydrodynamically Guided Hierarchical Self‐Assembly of Peptide–Protein Bioinks

Effective integration of molecular self‐assembly and additive manufacturing would provide a technological leap in bioprinting. This article reports on a biofabrication system based on the hydrodynamically guided co‐assembly of peptide amphiphiles (PAs) with naturally occurring biomolecules and proteins to generate hierarchical constructs with tuneable molecular composition and structural control. The system takes advantage of droplet‐on‐demand inkjet printing to exploit interfacial fluid forces and guide molecular self‐assembly into aligned or disordered nanofibers, hydrogel structures of different geometries and sizes, surface topographies, and higher‐ordered constructs bound by molecular diffusion. PAs are designed to co‐assemble during printing in cell diluent conditions with a range of extracellular matrix (ECM) proteins and biomolecules including fibronectin, collagen, keratin, elastin‐like proteins, and hyaluronic acid. Using combinations of these molecules, NIH‐3T3 and adipose derived stem cells are bioprinted within complex structures while exhibiting high cell viability (>88%). By integrating self‐assembly with 3D‐bioprinting, the study introduces a novel biofabrication platform capable of encapsulating and spatially distributing multiple cell types within tuneable pericellular environments. In this way, the work demonstrates the potential of the approach to generate complex bioactive scaffolds for applications such as tissue engineering, in vitro models, and drug screening.     

Surface Decoration of Peptide Nanoparticles Enables Efficient Therapy toward Osteoporosis and Diabetes

A versatile surface decoration strategy to efficiently encapsulate water-soluble peptides is developed. By assembling peptide molecules into nanoparticles, diverse physiochemical properties of these compacted molecules are equalized to the surface properties of nanoparticles. Primarily driven by the generic electrostatic attractions, the surface of as-prepared peptide nanoparticles is decorated with charged amino acids-grafted poly(lactic-co-glycolic acid). This adsorbed polymer layer versatilely blocks the phase transfer of peptide nanoparticles by increasing their affinity to the dispersed phase solvent molecules. Attributed to the ultrahigh encapsulation efficiencies (> 96%), the peptide mass fraction inside the obtained microcomposites is higher than 48%. The plasma calcium level has been efficiently reduced for ≈3 weeks with only one single injection of salmon calcitonin-encapsulated microcomposite in osteoporotic rats. Similarly, one single injection of exenatide-encapsulated microcomposites efficiently controls the glycemic level in type 2 diabetic rats for up to 3 weeks. Overall, the developed versatile surface decoration strategy efficiently encapsulates peptides and improves their pharmacokinetic features, regardless of the molecular structure of peptide cargos.     

Assemblies of Functional Peptides and Their Applications in Building Blocks for Biosensors

This feature article highlights our recent applications of functional peptide nanotubes, self‐assembled from short peptides with recognition elements, as building blocks to develop sensors. Peptide nanotubes with high aspect ratios are excellent building blocks for a directed assembly into device configurations, and their combined structures with nanometric diameters and micrometric lengths enables to bridge the “nanoworld” and the “microworld”. When the peptide‐nanotube‐based biosensors, which incorporate molecular recognition units, apply alternating current probes to detect impedance signals, the peptide nanotubes behave as excellent building blocks of the transducer for the detection of target analyes such as pathogens, cells, and heavey metal ions with high specificity. In some sensor configurations, the electric signal can be amplified by coupling them with ion‐specific mineralization via molecular recognition of peptides. In general the detection limit of peptide nanotube chips sensors is very low and the dynamic range of detection can be widened by improved device designs.     

Significant Enhancement of Proton Transport in Bioinspired Peptide Fibrils by Single Acidic or Basic Amino Acid Mutation

Bioinspired materials are extremely suitable for the development of biocompatible and environmentally friendly functional materials. Peptide‐based assemblies are remarkably attractive for such tasks, since they provide a simple way to fuse together functional and structural protein motifs in artificial materials. Motivated by this idea, it is shown here that the introduction of a single acidic, or basic, amino acid into the side chain of a heptameric self‐assembling peptide increases proton conduction in the resulting fibers by two orders of magnitude. This self‐doping effect is much more pronounced than the effect induced by the peptide's acidic and basic termini groups. Furthermore, the self‐doping process is found to be significantly more effective for acidic side chains than for basic ones due to both much more effective self‐doping process, resulting in an order of magnitude larger concentration of charge carriers for the acidic assemblies, and higher mobility of the formed charge carriers – almost threefolds in this case. This work facilitates the realization of unique bioinspired self‐assembled proton conducting materials that may find uses in the emerging bioprotonic technology. The presented design flexibility and, in particular, the ability to introduce both proton and proton holes further extend the usefulness of these materials.     

Effect of Heat Treatment Upon the Chemical Composition of Cottonseed Meal and Upon the Reactivity of Cottonseed Condensed Tannins

The effects of heat treatment on the chemical composition of cottonseed meal (CSM), with or without the addition of cottonseed hulls (containing condensed tannins; CT), and upon reactivity of the CT were studied. Heat was applied in a forced draught oven at 100°C for 2 h. Fluorodinitrobenzene (FDNB)-available lysine, free gossypol, extractable- and bound-CT concentrations, in vitro total nitrogen (N) solubility and the in vitro rumen degradation of the two major seed proteins (52 and 48 kDa) present in cottonseed kernel (which does not contain CT) were determined. The reactivity of CT was assessed by determining N solubility and rumen degradation of cottonseed kernel proteins in the presence or absence of polyethylene glycol (PEG; molecular weight (MW) 3500), which binds and inactivates CT. Heat treatment reduced the concentrations of free gossypol and FDNB-available lysine by small amounts, reduced measurable total CT content by 13%, reduced the solubility of total N, and reduced potential degradability of the 52 and 48 kDa cottonseed storage proteins by mixed rumen microorganisms. Addition of hulls further depressed solubility of total N and ruminal degradation of the two major storage proteins in cottonseed kernel. The action of PEG in vitro indicated that only part of the depression caused by hull addition could be explained by the presence of CT in the hulls, and that the effects of CT upon N solubility and potential degradability in heated CSM were similar to that in unheated CSM. Addition of hulls also substantially reduced FDNB-available lysine. In commercially produced materials, CSM from the Brisbane mill had a lower total CT content, lower N solubility and lower ruminal protein degradation rate than CSM from the Narrabri mill, but a similar level of FDNB-available lysine. Although application of heat inactivated 13% of the total CT, such that it could no longer be extracted and detected with butanol/HCl, it did not seem to change the overall effects produced by CT in reducing N solubility and protein degradation. The effect of hull addition in reducing available lysine has considerable relevance for feeding CSM to monogastric livestock. Interactions involving heat, hulls and CT need to be further studied.     

Relevance of AIF/CypA Lethal Pathway in SH-SY5Y Cells Treated with Staurosporine

The AIF/CypA complex exerts a lethal activity in several rodent models of acute brain injury. Upon formation, it translocates into the nucleus of cells receiving apoptotic stimuli, inducing chromatin condensation, DNA fragmentation, and cell death by a caspase-independent mechanism. Inhibition of this complex in a model of glutamate-induced cell death in HT-22 neuronal cells by an AIF peptide (AIF(370-394)) mimicking the binding site on CypA, restores cell survival and prevents brain injury in neonatal mice undergoing hypoxia-ischemia without apparent toxicity. Here, we explore the effects of the peptide on SH-SY5Y neuroblastoma cells stimulated with staurosporine (STS), a cellular model widely used to study Parkinson’s disease (PD). This will pave the way to understanding the role of the complex and the potential therapeutic efficacy of inhibitors in PD. We find that AIF(370-394) confers resistance to STS-induced apoptosis in SH-SY5Y cells similar to that observed with CypA silencing and that the peptide works on the AIF/CypA translocation pathway and not on caspases activation. These findings suggest that the AIF/CypA complex is a promising target for developing novel therapeutic strategies against PD.