Designed Peptide Assemblies for Efficient Gene Delivery.

The safe and efficient delivery of nucleic acids including DNA, mRNA, siRNA, and miRNA into targeted cells is critical for gene therapy. Currently, viral gene vectors are very popular, but they have potential toxicity and insecurity. Therefore, the development of nonviral vectors has attracted considerable research attention. Peptide assemblies are superior candidates for being used as gene vectors by having good biocompatibility, versatile molecular design, excellent assembly capacity, ease of modification, and stimuli responsivity. The de novo designed peptides not only can induce efficient condensation of nucleic acids into compacted nanoparticles and protect them from enzymatic digestion but also can effectively overcome biological barriers and improve gene delivery efficiency through targeted delivery, enhanced cellular uptake, improved endolysosomal escape, and nuclear importation. By having these merits, peptidic gene vectors are developing fast, showing outstanding advantages compared to liposome and polymer vectors. This Perspective focuses on peptidic gene delivery systems by emphasizing the molecular design strategies for meeting the criteria of gene condensation, protection from nuclease degradation, cellular uptake, endolysosomal escape, and so on. The new arising research area of peptide-based artificial viruses for gene and ribonucleoprotein delivery has also been reviewed. The challenges and future perspectives are put forward, aiming to provide a conclusive guide for the development of peptidic delivery systems to achieve efficient gene therapy.

Amino Acid-Mediated Synthesis of the ZIF-8 Nanozyme That Reproduces Both the Zinc-Coordinated Active Center and Hydrophobic Pocket of Natural Carbonic Anhydrase.

The zeolitic imidazolate framework-8 (ZIF-8) nanozyme has been synthesized using hydrophobic amino acid (AA) to regulate crystal growth. The as-synthesized ZIF-8 reproduces both the structural and functional properties of natural carbonic anhydrase (CA). Structurally, Zn2+/2-methylimidazole coordinated units mimic very well the active center of CA while the hydrophobic microdomains of the adsorbed AA simulate the CA hydrophobic pocket. Functionally, the nanozymes show excellent CA-like esterase activity by giving specific enzyme activity of 0.22 U mg-1 at 25 °C in the case of Val-ZIF-8. More strikingly, such nanozymes are superior to natural CA by having excellent hydrothermal stability, which can give highly enhanced esterase activity with increasing temperature. The specific enzyme activity of Val-ZIF-8 at 80 °C is about 25 times higher than that at 25 °C. In addition, AA-ZIF-8 also shows an excellent catalytic efficiency toward carbon dioxide (CO2) hydration. This study puts forward the important role of hydrophobic microdomains in biomimetic nanozymes for the first time and develops a facile and mild method for the synthesis of nanozymes with controlled morphology and size to achieve excellent catalytic efficiency.

Core-Shell Structures from the Coassembly of Lipoprotein-like Nanoparticles and Plasmid DNA for Gene Delivery.

We reported self-assembled core-shell nanoparticles (NPs) based on lipoprotein-like NPs and plasmid DNA (pDNA). Lipoprotein-like NPs were prepared using cholic acid (CA)-modified lipopeptides. We designed six different lipopeptides with different peptide segments to construct a series of NPs. It was proven that these NPs have different positive surface charges. These NPs could bind pDNA through electrostatic interaction to form core-shell complexes. The interactions between NPs and pDNA were systematically investigated. The number of NP charges determines the strength of the interaction between NPs and pDNA. Thus, various types of core-shell structures, such as loose and dense core-shell NPs, were found in this system. Cytotoxicity test confirmed that the carriers had no toxicity. We also proved that the core-shell structures have a good cell transfection effect. This study would expand the application of lipopeptide assemblies in the gene delivery field, which may lead to the development of peptide-based gene vectors for therapeutic application.

pH-Responsive Self-Assemblies from the Designed Folic Acid-Modified Peptide Drug for Dual-Targeting Delivery.

Targeting delivery is a promising technique for the therapy of cancers. A molecule FA-EEYSV-NH2, which consists of target recognition site folic acid (FA), dipeptide linker, and peptide drug, was designed as a novel anticancer prodrug. The molecules could self-assemble into nanoparticles at pH 7.0 and nanofibers at pH 5.0. By the aid of pH-responsiveness, the self-assemblies were used purposefully as targeted vehicles of self-delivery prodrugs. The results of cell toxicity and internalization assays have proved that the self-assemblies have good cancer cell selectivity. The selection was mainly attributed to the pH-responsive structure transition of self-assemblies and the FA active-targeting effect. We hope that our work could provide a useful strategy for finely tuning the properties and activities of peptide-based supramolecular nanomaterials, thus optimizing nanomedicines with enhanced performance.

Expression and characterization of catechol 1,2-dioxygenase from Oceanimonas marisflavi 102-Na3.

The gene of catechol 1, 2-dioxygenase was identified and cloned from the genome of Oceanimonas marisflavi 102-Na3. The protein was expressed in Escherichia coli BL21 (DE3) and purified to homogeneity of a dimer with molecular mass of 69.2 kDa. The enzyme was highly stable in pH 6.0-9.5 and below 45 °C and exhibited the maximum activity at pH 8.0 and 30 °C. Being the first characterized intradiol dioxygenase from marine bacteria Oceanimonas sp., the enzyme showed catalytic activity for catechol, 3-methylcatechol, 4-methylcatechol, 3-chlorocatechol, 4-chlorocatechol and pyrogallol. For catechol, Km and Vmax were 11.2 µM and 13.4 U/mg of protein, respectively. The enzyme also showed resistance to most of the metal ions, surfactants and organic solvents, being a promising biocatalyst for biodegradation of aromatic compounds in complex environments.

Peptide-Mediated Synthesis of Zeolitic Imidazolate Framework-8 with Controllable Morphology and Size.

Peptides with a sequence of Nap-Ix-GPLGLAG-R4-NH2 (x = 2, 4, and 6, shorted as I2R4, I4R4, and I6R4) were used as capping agents for the synthesis of zeolitic imidazolate framework-8 (ZIF-8) in water. Peptide addition can significantly inhibit the growth of ZIF-8 crystals. The shape and size of ZIF-8 crystals was related closely to the number of isoleucine (Ile, I) residues as well as concentration of the peptide. The shape of ZIF-8 crystals changes from rhomboid dodecahedron to truncated rhombic dodecahedron to cube with the decreasing number of isoleucine residues from six to two. At a peptide concentration of 1.0 mM, the morphology of ZIF-8 crystals was cubic, truncated rhombic dodecahedron, and typical rhombic dodecahedron in the cases of I2R4, I4R4, and I6R4, respectively. Also, the particle size can be regulated from ca. 1.7 µm to <100 nm by controlling the peptide concentration from 0 to 2.0 mM. This work develops a simple and green method for the synthesis of ZIF-8 crystals with controllable shape and size in water, which shows high potential for biomedical and biological applications.

Active Modulation of States of Prestress in Self-Assembled Short Peptide Gels.

Peptide hydrogels are excellent candidates for medical therapeutics due to their tuneable viscoelastic properties, however, in vivo they will be subject to various osmotic pressures, temperature changes, and biological co-solutes, which could alter their performance. Peptide hydrogels formed from the synthetic peptide I3K have a temperature-induced hardening of their shear modulus by a factor of 2. We show that the addition of uncross-linked poly( N-isopropylacrylamide) chains to the peptide gels increases the gels' temperature sensitivity by 3 orders of magnitude through the control of osmotic swelling and cross-linking. Using machine learning combined with single-molecule fluorescence microscopy, we measured the modulation of states of prestress in the gels on the level of single peptide fibers. A new self-consistent mixture model was developed to simultaneously quantify the energy and the length distributions of the states of prestress. Switching the temperature from 20 to 40 °C causes 6-fold increases in the number of states of prestress. At the higher temperature, many of the fibers experience constrained buckling with characteristic small wavelength oscillations in their curvature.

Reversible Thermoresponsive Peptide-PNIPAM Hydrogels for Controlled Drug Delivery.

Mixed thermoreversible gels were successfully fabricated by the addition of a thermosensitive polymer, poly(N-isopropylacrylamide) (PNIPAM), to fibrillar nanostructures self-assembled from a short peptide I3K. When the temperature was increased above the lower critical solution temperature of the PNIPAM, the molecules collapsed to form condensed globular particles, which acted as cross-links to connect different peptide nanofibrils and freeze their movements, resulting in the formation of a hydrogel. Since these processes were physically driven, such hydrogels could be reversibly switched between the sol and gel states as a function of temperature. As a model peptide, I3K was formulated with PNIPAM to produce a thermoreversible sol-gel system with a transition temperature of ∼33 °C, which is just below the body temperature. The antibacterial peptide of G(IIKK)3I-NH2 could be conveniently encapsulated in the hydrogel by the addition of the solution at lower temperatures in the sol phase and then increasing the temperature to be above 33 °C for gelation. The hydrogel gave a sustained and controlled linear release of G(IIKK)3I-NH2 over time. Using the peptide nanofibrils as three-dimensional scaffolds, such thermoresponsive hydrogels mimic the extracellular matrix and could potentially be used as injectable hydrogels for minimally invasive drug delivery or tissue engineering.

Self-Assembly of Short Elastin-like Amphiphilic Peptides: Effects of Temperature, Molecular Hydrophobicity and Charge Distribution.

A novel type of self-assembling peptides has been developed by introducing the basic elastomeric ß-turn units of elastin protein into the amphiphilic peptide molecules. The self-assembly behaviors of such peptides are affected by the overall molecular hydrophobicity, charge distribution and temperature. The molecules with higher hydrophobicity exhibit better self-assembling capability to form long fibrillar nanostructures. For some peptides, the temperature increase can not only promote the self-assembly process but also change the self-assembly routes. The self-assembly of the peptides with two charges centralized on one terminal show higher dependence on temperature than the peptides with two charges distributed separately on the two terminals. The study probes into the self-assembly behaviors of short elastin-like peptides and is of great help for developing novel self-assembling peptides with thermo sensitivity.

Dual-Responsive Viscoelastic Lyotropic Liquid Crystal Fluids to Control the Diffusion of Hydrophilic and Hydrophobic Molecules.

A smart lyotropic liquid crystal (LLC) system was prepared to control the diffusion rate of hydrophilic and hydrophobic molecules. The LLC system is composed of a nonionic surfactant (tetraethylene glycol monododecylether; C12 EO4 ) and an anionic azobenzene surfactant (Azo-surfactant). C12 EO4 was the main component of the LLC system. The Azo-surfactant, which can undergo photo-isomerization, played the role of trigger in this system. LLC gels formed in a solution comprised of Azo-surfactant (10 mm) and C12 EO4 (300 mm). The LLC gels became broken when more Azo-surfactant was added (e.g., up to 15 mm) and the viscoelasticity was lost. Surprisingly, when we used UV light to irradiate the 300 mm C12 EO4 /15 mm Azo-surfactant sample, the gel was recovered and high viscoelasticity was observed. However, under visible-light irradiation, the gel became broken again. The gel formation could also be triggered by heating the sample. On heating the 300 mm C12 EO4 /15 mm Azo-surfactant sample, the system thickened to a point at which typical gel behavior was registered. When the sample was cooled, the gel broke again. The LLC could be used for controlled release of hydrophilic and hydrophobic molecules, and could be considered as a versatile vehicle for the delivery of actives in systems of practical importance.

Self-assembled two-dimensional thermoresponsive microgel arrays for cell growth/detachment control.

Monodisperse poly(N-isopropylacrylamide-styrene) (PNIPAAmSt) microgels with different St/NIPAAm ratios have been synthesized via a one-step surfactant-free emulsion polymerization process. The resulting microgel dispersions were used to fabricate 2D arrays on the surface of silicon wafers/glass coverslips through dip coating. The thermal responsiveness of the PNIPAAmSt microgel arrays was examined by spectroscopic ellipsometry and the results unraveled that the thermoresponsive behavior of the arrays was highly consistent with the microgels dispersed in the bulk, showing high dependence on the content of styrene. The structure of the films varied from nonclose-packed 2D arrays to close-packed 2D arrays, depending on both properties of the microgels and array fabrication conditions. When the weight ratio of styrene was below 40%, the microgel arrays demonstrated effective control for cell growth and detachment across their volume phase transition temperatures (around 28 °C). The extent of swelling of the microgels was the key factor to determine whether the cells could detach from the film easily. For the rather close-packed 2D arrays prepared by the same kind of PNIPAAmSt microgels, the gaps between microgel particles showed no obvious effect on the rate of cell detachment.

Recent Development of Nanomaterials for Chemical Engineering.

There has been an explosive growth in research on nanomaterials since the late 1980s and early 1990s [...].

Bicontinuous Interfacially Jammed Emulsion Gels (Bijels): Preparation, Control Strategies, and Derived Porous Materials.

Bicontinuous interfacially jammed emulsion gels, also known as Bijels, are a new type of soft condensed matter. Over the last decade, Bijels have attracted considerable attention because of their unique morphology, property, and broad application prospects. In the present review, we summarize the preparation methods and main control strategies of Bijels, focusing on the research progress and application of Bijels as templates for porous materials preparation in recent years. The potential future directions and applications of Bijels are also envisaged.

Thermoresponsive microgel films for harvesting cells and cell sheets.

This work reports the formation of thermoresponsive poly(N-isopropylacrylamide-co-styrene) (PNIPAAmSt) microgel films and their use for cell growth and detachment via temperature stimuli. Thermoresponsive surface films can be conveniently produced by spin-coating or drop-coating of PNIPAAmSt microgel dispersions onto substrates such as glass coverslips, cell culture plates, and flasks, making this technique widely accessible. The thickness, stability, and reversibility of the PNIPAAmSt films coated on silicon wafers with respect to temperature switching were examined by spectroscopic ellipsometry (SE) and atomic force microscopy (AFM). The results unraveled the direct link between thermoreversibility and changes in film thickness and surface morphology, showing reversible hydration and dehydration. Under different coating conditions, well-packed microgel monolayers could be utilized for effective cell recovery and harvesting. Furthermore, cell adhesion and detachment processes were reversible and there was no sign of loss of cell viability during repeated surface attachment, growth, and detachment, showing a mild interaction between cells and thermoresponsive surface. More importantly, there was little deterioration of the packing of the thermoresponsive films or any major loss of microgel particles during reuse, indicating their robustness. These PNIPAAmSt microgel films thus open up a convenient interfacial platform for cell and cell sheet harvesting while avoiding the damage of enzymatic cleavage.

Development of Functional Nanomaterials for Applications in Chemical Engineering.

Nanomaterials are materials with particle sizes of less than 100 nm in at least one of their dimensions [...].

Synthesis and Evaluation of Peptide-Manganese Dioxide Nanocomposites as Adsorbents for the Removal of Strontium Ions.

In this study, a novel organic-inorganic hybrid material IIGK@MnO2 (2-naphthalenemethyl-isoleucine-isoleucine-glycine-lysine@manganese dioxide) was designed as a novel adsorbent for the removal of strontium ions (Sr2+). The morphology and structure of IIGK@MnO2 were characterized using TEM, AFM, XRD, and XPS. The results indicate that the large specific surface area and abundant negative surface charges of IIGK@MnO2 make its surface rich in active adsorption sites for Sr2+ adsorption. As expected, IIGK@MnO2 exhibited excellent adsorbing performance for Sr2+. According to the adsorption results, the interaction between Sr2+ and IIGK@MnO2 can be fitted with the Langmuir isotherm and pseudo-second-order equation. Moreover, leaching and desorption experiments were conducted to assess the recycling capacity, demonstrating significant reusability of IIGK@MnO2.

Cancer-Responsive Multifunctional Nanoplatform Based on Peptide Self-Assembly for Highly Efficient Combined Cancer Therapy by Alleviating Hypoxia and Improving the Immunosuppressive Microenvironment.

Hypoxia, as a main feature of the tumor microenvironment, has greatly limited the efficacy of photodynamic therapy (PDT), as well as its clinical application. Here, a multifunctional composite nanoplatform, the peptide/Ce6/MnO2 nanocomposite (RKCM), has been constructed to alleviate tumor hypoxia and increase the efficacy of PDT using rationally designed peptide fibrils to encapsulate chlorin e6 (Ce6) inside and to mineralize MnO2 nanoparticles on the surface. As a result, RKCM significantly improved the PDT efficacy by increasing reactive oxygen species (ROS) generation, decreasing tumor cell viability, and inhibiting tumor growth and metastasis. Besides, decreased HIF-1α expression and increased immune-activated cell infiltration were also observed in RKCM/laser treatment xenograft. Mechanically, (1) Ce6 can induce singlet oxygen (1O2) generation under laser irradiation to give photodynamic therapy (PDT); (2) MnO2 can react with H2O2 in situ to supply additional O2 to alleviate tumor hypoxia; and (3) the released Mn2+ ions can induce a Fenton-like reaction to generate •OH for chemical dynamic therapy (CDT). Moreover, RKCM/laser treatment also presented with an abscopal effect to block the occurrence of lung metastasis by remolding the pre-metastasis immune microenvironment. With these several aspects working together, the peptide/Ce6/MnO2 nanoplatform can achieve highly efficient tumor therapy. Such a strategy based on peptide self-assembly provides a promising way to rationally design a cancer-responsive multifunctional nanoplatform for highly efficient combined cancer therapy by alleviating hypoxia and improving the immune microenvironment.

Gold nanoparticles inhibit tumor growth via targeting the Warburg effect in a c-Myc-dependent way.

Gold nanoparticles (AuNPs) are prospective tools for nano-based medicine that can directly target cellular biological processes to influence cell fate and function. Studies have revealed the essential role of AuNPs in metabolic remodeling for macrophage polarization. Nevertheless, as a hallmark of cancer cells, metabolic changes in tumor cells in response to AuNPs have not yet been reported. In the present study, polymer- and folate-conjugated AuNPs with satisfactory biocompatibility and tumor-targeting activity were synthesized to investigate their underlying roles in tumor metabolism. Tumor cells were significantly suppressed by AuNPs in vitro and in vivo, with little cytotoxicity in non-tumor cells. Subcellular localization showed that AuNPs localized in the mitochondria of tumor cells and impaired their structure and function, leading to excessive oxidative stress and mitochondrial apoptosis. Metabolic stress, with decreased glycolysis and insufficient nutrients, was also caused by AuNPs exposure in tumor cells. Mechanistically, the key enzymes (GLUT1 and HK2) for glycolysis modulation were remarkably reduced by AuNPs in a c-Myc-dependent manner. The present study demonstrated a new mechanism for AuNPs in the inhibition of tumor growth, that is, via directly targeting glycolysis and depriving energy. These findings provide new strategies for the design of nano-based medicines and anti-glycolytic therapeutics to inhibit the development of malignant tumors. STATEMENT OF SIGNIFICANCE: Gold nanoparticles (AuNPs) have acquired ever-increasing interest for applications in cancer treatment and diagnosis due to their high biosafety and facile surface modification. Recent studies have shown that AuNPs can work as active agents to directly target the cellular processes and harbor antitumor properties, while the underlying mechanisms remain largely unknown. From the present findings, the stabilized AuNPs showed direct inhibition effects on tumor growth by glycolysis inhibition and energy deprivation. These results provide new insights of AuNPs for tumor treatments, which will further contribute to the development of promising nano-based medicines and anti-glycolytic therapies.

Characterization of photosystem I from spinach: effect of solution pH.

Our previous work has demonstrated the isolation of photosystem I (PSI) from spinach using ultrafiltration with a final purity of 84%. In order to get a higher purity of PSI and more importantly to develop a practical bioseparation process, key physiochemical properties of PSI and their dependence on operational parameters must be assessed. In this study, the effect of solution pH, one of the most important operating parameters for membrane process, on the property of PSI was examined. Following the isolation of crude PSI from spinach using n-dodecyl-beta-D: -maltoside as detergent, the isoelectric point, aggregation size, zeta potential, low-temperature fluorescence, atomic force microscopy imaging, secondary structure, and thermal stability were determined. Solution pH was found to have a significant effect on the activity, aggregation size and thermal stability of PSI. The results also suggested that the activity of PSI was related to its aggregation size.

Application of Peptides in Construction of Nonviral Vectors for Gene Delivery.

Gene therapy, which aims to cure diseases by knocking out, editing, correcting or compensating abnormal genes, provides new strategies for the treatment of tumors, genetic diseases and other diseases that are closely related to human gene abnormalities. In order to deliver genes efficiently to abnormal sites in vivo to achieve therapeutic effects, a variety of gene vectors have been designed. Among them, peptide-based vectors show superior advantages because of their ease of design, perfect biocompatibility and safety. Rationally designed peptides can carry nucleic acids into cells to perform therapeutic effects by overcoming a series of biological barriers including cellular uptake, endosomal escape, nuclear entrance and so on. Moreover, peptides can also be incorporated into other delivery systems as functional segments. In this review, we referred to the biological barriers for gene delivery in vivo and discussed several kinds of peptide-based nonviral gene vectors developed for overcoming these barriers. These vectors can deliver different types of genetic materials into targeted cells/tissues individually or in combination by having specific structure-function relationships. Based on the general review of peptide-based gene delivery systems, the current challenges and future perspectives in development of peptidic nonviral vectors for clinical applications were also put forward, with the aim of providing guidance towards the rational design and development of such systems.