Shengjie Wang

Tuning the Self-Assembly of Short Peptides via Sequence Variations

Peptide self-assembly is of direct relevance to protein science and bionanotechnology, but the underlying mechanism is still poorly understood. Here, we demonstrate the distinct roles of the noncovalent interactions and their impact on nanostructural templating using carefully designed hexapeptides, I2K2I2, I4K2, and KI4K. These simple variations in sequence led to drastic changes in final self-assembled structures. β-sheet hydrogen bonding was found to favor the formation of one-dimensional nanostructures, such as nanofibrils from I4K2 and nanotubes from KI4K, but the lack of evident β-sheet hydrogen bonding in the case of I2K2I2 led to no nanostructure formed. The lateral stacking and twisting of the β-sheets were well-linked to the hydrophobic and electrostatic interactions between amino acid side chains and their interplay. For I4K2, the electrostatic repulsion acted to reduce the hydrophobic attraction between β-sheets, leading to their limited lateral stacking and more twisting, and final fibrillar structures; in contrast, the repulsive force had little influence in the case of KI4K, resulting in wide ribbons that eventually developed into nanotubes. The fibrillar and tubular features were demonstrated by a combination of cryogenic transmission electron microscopy (cryo-TEM), negative-stain transmission electron microscopy (TEM), and small-angle neutron scattering (SANS). SANS also provided structural information at shorter scale lengths. All atom molecular dynamics (MD) simulations were used to suggest possible molecular arrangements within the β-sheets at the very early stage of self-assembly.

Rapid microwave-assisted synthesis of ultra-bright fluorescent carbon dots for live cell staining, cell-specific targeting and in vivo imaging

Highly fluorescent carbon dots (CDs) with quantum yields up to 96% were rapidly synthesized within 5 min via microwave irradiation with controllable temperature. Multifunctional bioimaging including live cell staining, cell-specific targeting and in vivo imaging were further demonstrated by using high quality and low cost CDs as contrast agents.

Visible and Near-Infrared Dual-Emission Carbogenic Small Molecular Complex with High RNA Selectivity and Renal Clearance for Nucleolus and Tumor Imaging

Fluorescence imaging requires bioselective, sensitive, nontoxic molecular probes to detect the precise location of lesions for fundamental research and clinical applications. Typical inorganic semiconductor nanomaterials with large sizes (>10 nm) can offer high-quality fluorescence imaging due to their fascinating optical properties but are limited to low selectivity as well as slow clearance pathway. We here report an N- and O-rich carbogenic small molecular complex (SMC, MW < 1000 Da) that exhibits high quantum yield (up to 80%), nucleic acid-binding enhanced excitation-dependent fluorescence (EDF), and a near-infrared (NIR) emission peaked at 850 nm with an ultralarge Stokes shift (∼500 nm). SMCs show strong rRNA affinity, and the resulting EDF enhancement allows multicolor visualization of nucleoli in cells for clear statistics. Furthermore, SMCs can be efficiently accumulated in tumor in vivo after injection into tumor-bearing mice. The NIR emission affords high signal/noise ratio imaging for delineating the true extent of tumor. Importantly, about 80% of injected SMCs can be rapidly excreted from the body in 24 h. No appreciable toxicological responses were observed up to 30 days by hematological, biochemical, and pathological examinations. SMCs have great potential as a promising nucleolus- and tumor-specific agent for medical diagnoses and biomedical research.

Synthesis of 1D Silica Nanostructures with Controllable Sizes Based on Short Anionic Peptide Self-Assembly

Artificial synthesis of silica under benign conditions is usually achieved by using cationic organic matrices as templates while the anionic analogues have not received enough consideration, albeit they are also functioning in biosilica formation. In this work, we report the design and self-assembly of an anionic peptide amphiphile (I3E) and the use of its self-assemblies as templates to synthesize 1D silica nanostructures with tunable sizes. We show that short I3E readily formed long nanofibrils in aqueous solution via a hierarchical self-assembly process. By using APTES and TEOS as silica precursors, we found that the I3E nanofibrils templated the production of silica nanotubes with a wide size distribution, in which the silica size regulation was achieved by tuning the interactions among the peptide template and silicon species. These results clearly illustrate a facile method for generating silica nanomaterials based on anionic matrices.

Peptide-templated synthesis of branched MnO2 nanowires with improved electrochemical performances

Although many nanomaterials have been prepared in vitro by mimicking biomineralization, the biomimetic synthesis of hybrids with both well-ordered nanostructures and specific functions is still in its infancy. A short designed peptide amphiphile I3K can form uniform and stable nanofibers in aqueous solution, with a surface enriched in cationic lysine residue. In the present study, we have demonstrated that the peptide nanofibers could direct the synthesis of MnO2 nanowires under mild conditions. By varying the concentration of manganese precursors (KMnO4 and Mn(NO3)2), uniform branched MnO2/peptide hybrid nanowires with high porosity and a large specific surface area were obtained. The well-defined MnO2 hybrid nanowires showed significantly improved electrochemical supercapacitive properties relative to compact MnO2 nanowires and urchin-like MnO2 spheres. Their specific capacitance could attain a higher value of 421 F g−1 and retained about 93% of the initial capacitance after 2500 cycles at a scan rate of 5 mV s−1, and remained little changed during the process of progressively varying the current density. Furthermore, the electrode prepared from the uniform MnO2 hybrid nanowires showed an excellent reversibility and a reasonably high-rate capability during the charge/discharge process. Such a study provides a new methodology to prepare functional MnO2 nanostructures under mild conditions that can be used in electrochemical energy storage.

Deep-Red Fluorescent Gold Nanoclusters for Nucleoli Staining: Real-Time Monitoring of the Nucleolar Dynamics in Reverse Transformation of Malignant Cells

Nucleoli are important subnuclear structures inside cells. We report novel fluorescent gold nanoclusters (K-AuNCs) that are able to stain the nucleoli selectively and make it possible to explore the nucleolar morphology with fluorescence imaging technique. This novel probe is prepared through an easy synthesis method by employing a tripeptide (Lys-Cys-Lys) as the surface ligand. The properties, including deep-red fluorescence emission (680 nm), large Stocks shift, broad excitation band, low cytotoxicity, and good photostability, endow this probe with potential for bioanalytical applications. Because of their small size and their positively charged surface, K-AuNCs are able to accumulate efficiently at the nucleolar regions and provide precise morphological information. K-AuNCs are also used to monitor the nucleolar dynamics along the reverse-transformation process of malignant cells, induced by the agonist of protein A, 8-chloro-cyclic adenosine monophosphate. This gives a novel approach for investigating the working mechanism of antitumor drugs.

Fabrication of Patterned Thermoresponsive Microgel Strips on Cell-Adherent Background and Their Application for Cell Sheet Recovery

Interfaces between materials and cells play a critical role in cell biomedical applications. Here, a simple, robust, and cost-effective method is developed to fabricate patterned thermoresponsive poly(N-isopropylacrylamide-co-styrene) microgel strips on a polyethyleneimine-precoated, non-thermoresponsive cell-adherent glass coverslip. The aim is to investigate whether cell sheets could be harvested from these cell-adherent surfaces patterned with thermoresponsive strips comprised of the microgels. We hypothesize that if the cell-to-cell interaction is strong enough to retain the whole cell sheet from disintegration, the cell segments growing on the thermoresponsive strips may drag the cell segments growing on the cell-adherent gaps to detach, ending with a whole freestanding and transferable cell sheet. Critical value concerning the width of the thermoresponsive strip and its ratio to the non-thermoresponsive gap may exist for cell sheet recovery from this type of surface pattern. To obtain this critical value, a series of strip patterns with various widths of thermoresponsive strip and non-thermoresponsive gap were prepared using negative microcontact printing technology, with COS7 fibroblast cells being used to test the growth and detachment. The results unraveled that COS7 cells preferentially attached and proliferated on the cell-adherent, non-thermoresponsive gaps to form patterned cell layers and that they subsequently proliferated to cover the microgel strips to form a confluent cell layer. Intact COS7 cell sheets could be recovered when the width of the thermoresponsive strip is no smaller than that of the non-thermoresponsive gap. Other cells such as HeLa, NIH3T3, 293E, and L929 could grow similarly; that is, they showed initial preference to the non-thermoresponsive gaps and then migrated to cover the entire patterned surface. However, it was difficult to detach them as cell sheets due to the weak interactions within the cell layers formed. In contrast, when COS7 and HeLa cells were cultured successively, they formed the cocultured cell layer that could be detached together. These freestanding patterned cell sheets could lead to the development of more elaborate tumor models for drug targeting and interrogation.

High-Density Super-Resolution Localization Imaging with Blinking Carbon Dots

Molecular fluorescence blinking provides a simple and attractive way to achieve super-resolution localization via conventional fluorescence microscopy. However, success in super-resolution imaging relies heavily on their blinking characteristics. We here report easily prepared and photostable nanoparticles, carbon dots (CDs), with desirable fluorescence blinking for high-density super-resolution imaging. The CDs exhibit a low duty cycle (∼0.003) and high photon output (∼8000) per switching event, as well as show much higher resistance to photobleaching than Alexa 647 or Cy5 typically used in single molecule localization microscopy. The stable blinking of CDs allows to perform high-density localization imaging at a resolution of 25 nm by sequentially recording the particle positions. The CD-based super-resolution imaging is further demonstrated by rendering CD-stained tubular peptide self-assemblies, CD-packed clusters with well-defined patterns, and CD-stained microtubules in a cell. Furthermore, this method has been validated as a valuable tool to detect the clustering and distribution of protein receptors in the plasma membrane that are not discerned with normal fluorescence imaging.

Virus-like supramolecular assemblies formed by cooperation of base pairing interaction and peptidic association

A peptide nucleic acid (PNA)-peptide conjugated molecule, T′3(AKAE)2, was designed to have both a PNA segment for oligonucleotide binding and an ionic self-complementary peptide sequence for self-association. T′3(AKAE)2 could co-assemble with oligoadenines (d(A)x) to form virus-like supramolecular structures whose morphology showed dependence on the chain length and rigidity of the d(A)x molecules. Smaller nanospheres with diameters of 13.0±2.0 nm were produced in the case of d(A)6. Wormlike aggregates with lengths of 20–50 nm and diameters of 15.0±2.5 nm were found in the cases of d(A)12, d(A)18, d(A)24 and d(A)30. And larger spherical aggregates with diameters of 18±5 nm came into presence in the cases of d(A)36 and d(A)42. These nanostructures were suggested to be formed under a cooperative effect of base pair recognition and peptidic association. The study provides insights into the programmed assembly of a multi-components system as well as control of the size and shape of the co-assembled structures, which is of great significance in developing gene/drug delivery systems.